WO2023197003A2 - Making cheese with soy proteins - Google Patents

Abstract

Compositions, methods and systems for making cheese using a soy ingredient having one or more soy proteins. In some instances, the soy protein is a storage protein comprising at least one of 7s (e.g., 0-conglycinin) or I ls (e.g., glycinin). In some cases, the ratio of 7s/l Is is lower than a naturally occurring ratio. In some aspects, current disclosure provides a new composition of milk augmented with soy proteins that minimizes the negative effects of soy on coagulation of casein micelles. In some instances, the soy ingredient is modified to decrease the ratio of 7s (e.g., 0- conglycinin) to 1 Is (e.g., glycinin), resulting in improved coagulation of casein micelles and cheese production.

Description

MAKING CHEESE WITH SOY PROTEINS

CROSS-REFERENCE

[0001] This application claims the benefit of U.S. Provisional Application No. 63/329,351, filed on April 8, 2022, U.S. Provisional Application No. 63/351,182, filed on June 10, 2022, U.S. Provisional Application No. 63/376,223, filed on September 19, 2022, and U.S. Provisional Application No. 63/444,189, filed on February 8, 2023, all of which are incorporated herein by reference in its entirety.

INCORPORATION BY REFERENCE

[0002] All publications, patents, and patent applications mentioned in this specification are herein incorporated by reference to the same extent as if each individual publication, patent, or patent application was specifically and individually indicated to be incorporated by reference.

SEQUENCE LISTING

[0003] The instant application contains a Sequence Listing which has been submitted electronically in XML format and is hereby incorporated by reference in its entirety. Said XML copy, created on April 7, 2023, is named 62126-706_601_SL.xml and is 22,977 bytes in size.

BACKGROUND

[0004] Soybeans are high in protein and are grown in large quantities throughout the world, making soy protein an inexpensive source of protein. Substitution of animal and microbial proteins with soy proteins can result in economic benefit for food producers. Unfortunately, soy components negatively impact the functionality of casein micelles resulting in poor coagulation, a loose curd, and lengthened clotting time during cheese production.

SUMMARY

[0005] The current disclosure provides compositions, methods and systems for making cheese using a soy ingredient having one or more soy proteins. In some instances, the soy ingredient is a seed storage protein comprising at least one of 7s (e.g., P-conglycinin) and 11s (e.g., glycinin), wherein the weight or molar ratio of 7s/l Is is less than a naturally occurring weight or molar ratio. In some aspects, the current disclosure provides a new composition of milk augmented with soy proteins that minimizes the negative effects of soy on coagulation of casein micelles. In some instances, the soy ingredient is modified to change the ratio of conglycinin to glycinin, resulting in improved coagulation of casein micelles and cheese production. [0006] In some aspects, the current disclosure provides a composition, comprising casein micelles; and a soy ingredient comprising at least one of 7s (e.g., P-conglycinin) or 11s (e.g., glycinin). In some cases, the composition is milk, cheese, or a curd. In some cases, the soy ingredient comprises only one of 7s (e.g., P-conglycinin) or 1 Is (e.g., glycinin), but not both. In some cases, the soy ingredient comprises both 7s (e.g., P-conglycinin) and 11s (e.g., glycinin), and the weight or molar ratio of 7s/l Is is lower than a naturally occurring ratio. In some instances, a decreased ratio of 7s/l Is is achieved by decreasing the amount of 7s (e.g., P- conglycinin) in the soy ingredient, for example, by using RNAi or CRISPR-Cas9 to knock out or knock down the gene expression of a plant protein, in this instance, 7s (e.g., P-conglycinin). In some instances, a decreased weight ratio of 7s/l Is is achieved by increasing the amount of 1 Is (e.g., glycinin) in the soy ingredient, for example, by overexpressing a l ls protein (e.g., glycinin) [0007] In some cases, the soy ingredient comprises both 7s (e.g., P-conglycinin) and I ls (e.g., glycinin), and the weight ratio of 11 s/7s is lower than a naturally occurring ratio. In some instances, a decreased ratio of 11 s/7s is achieved by decreasing the amount of 1 Is (e.g., glycinin) in the soy ingredient, for example, by using RNAi or CRISPR-Cas9 to knock out or knock down the gene expression of a plant protein, in this instance, I ls (e.g., glycinin). In some instances, a decreased weight ratio of 11 s/7s is achieved by increasing the amount of 7s (e.g., P-conglycinin) in the soy ingredient, for example, by overexpressing a 7s protein.

[0008] In some instances, at least one of the soy ingredients is recombinant or genetically modified. In some cases, 7s (e.g., P-conglycinin) is recombinant or genetically modified. In some cases, I ls (e.g., glycinin) is recombinant or genetically modified.

[0009] In some cases, the naturally occurring weight ratio of 7s/l Is is between 0.5 and 1.3. In some cases, the naturally occurring weight ratio of 7s/l Is is 0.5. In some cases, the naturally occurring weight ratio of 7s/l Is is 0.6. In some cases, the naturally occurring weight ratio of 7s/l Is is 0.7. In some cases, the naturally occurring weight ratio of 7s/l Is is 0.8. In some cases, the naturally occurring weight ratio of 7s/l Is is 0.75. In some cases, the naturally occurring weight ratio of 7s/l 1 s is 0.9. In some cases, the naturally occurring weight ratio of 7s/l 1 s is 1. In some cases, the naturally occurring weight ratio of 7s/l 1 s is 1.1. In some cases, the naturally occurring weight ratio of 7s/l 1 s is 1.2. In some cases, the naturally occurring weight ratio of 7s/l ls is 1.3.

[0010] In some cases, the weight ratio of 7s/l Is is lower than 0.5. In some cases, the weight ratio of 7s/l Is is lower than 0.45. In some cases, the weight ratio of 7s/l Is is lower than 0.4. In some cases, the weight ratio of 7s/l Is is lower than 0.35. In some cases, the weight ratio of 7s/l Is is lower than 0.3. In some cases, the weight ratio of 7s/l Is is lower than 0.25. In some cases, the weight ratio of 7s/l Is is lower than 0.2. In some cases, the weight ratio of 7s/l Is is lower than 0.15. In some cases, the weight ratio of 7s/l Is is lower than 0.1. In some cases, the weight ratio of 7s/l Is is lower than 0.05. In some cases, the weight ratio of 7s/l Is is lower than 0.01. In some cases, the weight ratio of 7s/l Is is lower than 0.001. In some cases, the weight ratio of 7s/l Is is lower than 0.0001. In some cases, the weight ratio of 7s/l Is is lower than 0.00001. In some cases, the weight ratio of 7s/l Is is lower than 0.000001. In some cases, the weight ratio of 7s/l 1 s is lower than 0.0000001. In some cases, the weight ratio of 7s/l Is is 0 or close to 0, wherein the soy ingredient comprises a trace amount of 7s (e.g., P-conglycinin), or does not comprise a detectable amount of 7s (e.g., P-conglycinin).

[0011] Some aspects of the current disclosure provide compositions comprising 7s or I ls. In some cases, the weight ratio of 11 s/7s is lower than 0.77. In some cases, the weight ratio of 11 s/7s is lower than 0.7. In some cases, the weight ratio of 11 s/7s is lower than 0.6. In some cases, the weight ratio of 11 s/7s is lower than 0.5. In some cases, the weight ratio of 11 s/7s is lower than 0.45. In some cases, the weight ratio of 11 s/7s is lower than 0.4. In some cases, the weight ratio of 11 s/7s is lower than 0.35. In some cases, the weight ratio of 11 s/7s is lower than 0.3. In some cases, the weight ratio of 11 s/7s is lower than 0.25. In some cases, the weight ratio of 11 s/7s is lower than 0.2. In some cases, the weight ratio of 11 s/7s is lower than 0.15. In some cases, the weight ratio of 11 s/7s is lower than 0.1. In some cases, the weight ratio of 11 s/7s is lower than 0.05. In some cases, the weight ratio of 11 s/7s is lower than 0.01. In some cases, the weight ratio of 11 s/7s is lower than 0.001. In some cases, the weight ratio of 11 s/7s is lower than 0.0001. In some cases, the weight ratio of 11 s/7s is lower than 0.00001. In some cases, the weight ratio of 11 s/7s is lower than 0.000001. In some cases, the weight ratio of 11 s/7s is lower than 0.0000001. In some cases, the weight ratio of 11 s/7s is close to 0, wherein the soy ingredient comprises only trace amount of 1 Is (e.g., glycinin), or does not comprise a detectable amount of 1 Is (e.g., glycinin).

[0012] Some aspects of the current disclosure provide compositions comprising casein micelles; and a soy ingredient comprising at least one of 7s or 1 Is, wherein the weight or molar ratio of the 7s and the I ls (7s/l Is) is higher than a naturally occurring ratio. In some cases, the naturally occurring weight ratio of the 7s and the I ls (7s/l Is) is between 0.5-1.3. In some cases, the ratio of the 7s and the I ls (7s/l Is) in the composition is higher than 1.3. In some cases, the ratio of the 7s and the I ls (7s/l Is) is higher than 2. In some cases, the ratio of the 7s and the I ls (7s/l Is) is higher than 5. In some cases, the ratio of the 7s and the I ls (7s/l Is) is higher than 10. In some cases, the ratio of the 7s and the I ls (7s/l Is) is higher than 20. In some cases, the ratio of the 7s and the I ls (7s/l Is) is higher than 50. In some cases, the ratio of the 7s and the I ls (7s/l Is) is higher than 100. In some cases, the ratio of the 7s and the I ls (7s/l Is) is higher than 1000. In some cases, the casein micelle comprises recombinant casein proteins. [0013] In some cases, the disclosed composition comprises 7s and I ls, where the molar ratio of 7s/l Is is lower than 0.5. In some cases, the molar ratio of 7s/l Is is lower than 0.45. In some cases, the molar ratio of 7s/l Is is lower than 0.4. In some cases, the molar ratio of 7s/l Is is lower than 0.35. In some cases, the molar ratio of 7s/l Is is lower than 0.3. In some cases, the molar ratio of 7s/l Is is lower than 0.25. In some cases, the molar ratio of 7s/l Is is lower than 0.2. In some cases, the molar ratio of 7s/l Is is lower than 0.15. In some cases, the molar ratio of 7s/l Is is lower than 0.1. In some cases, the molar ratio of 7s/l Is is lower than 0.05. In some cases, the molar ratio of 7s/l Is is lower than 0.01. In some cases, the molar ratio of 7s/l Is is lower than 0.001. In some cases, the molar ratio of 7s/l Is is lower than 0.0001. In some cases, the molar ratio of 7s/l Is is lower than 0.00001. In some cases, the molar ratio of 7s/l 1 s is lower than 0.000001. In some cases, the molar ratio of 7s/l 1 s is lower than 0.0000001. In some cases, the molar ratio of 7s/l Is is 0 or close to 0, wherein the soy ingredient comprises a trace amount of 7s (e.g., P-conglycinin), or does not comprise a detectable amount of 7s (e.g., P-conglycinin). [0014] In some cases, the disclosed composition comprises 7s and I ls, where molar ratio of 11 s/7s is lower than 0.77. In some cases, the molar ratio of 11 s/7s is lower than 0.7. In some cases, the molar ratio of 11 s/7s is lower than 0.6. In some cases, the molar ratio of 11 s/7s is lower than 0.5. In some cases, the molar ratio of 11 s/7s is lower than 0.45. In some cases, the molar ratio of 11 s/7s is lower than 0.4. In some cases, the molar ratio of 11 s/7s is lower than 0.35. In some cases, the molar ratio of 11 s/7s is lower than 0.3. In some cases, the molar ratio of 11 s/7s is lower than 0.25. In some cases, the molar ratio of 11 s/7s is lower than 0.2. In some cases, the molar ratio of 11 s/7s is lower than 0.15. In some cases, the molar ratio of 11 s/7s is lower than 0.1. In some cases, the molar ratio of 1 ls/7s is lower than 0.05. In some cases, the molar ratio of 11 s/7s is lower than 0.01. In some cases, the molar ratio of 11 s/7s is lower than 0.001. In some cases, the molar ratio of 11 s/7s is lower than 0.0001. In some cases, the molar ratio of 11 s/7s is lower than 0.00001. In some cases, the molar ratio of 11 s/7s is lower than 0.000001. In some cases, the molar ratio of 11 s/7s is lower than 0.0000001. In some cases, the molar ratio of 11 s/7s is close to 0, wherein the soy ingredient comprises only trace amount of 1 Is (e.g., glycinin), or does not comprise a detectable amount of 1 Is (e.g., glycinin).

[0015] In some cases, the soy ingredient is in the form of soy protein isolate. In some cases, the soy ingredient is in the form of soymilk. It is contemplated that disclosed ratios, weight percentages of different proteins are present in a dairy product, milk, cheese, cheese curd, or an intermediate product made during the process of making a dairy product (e.g., cheese). In some cases, the source of the casein micelles is from an animal, for example a mammal, for example, a human. In some cases, the mammal is a ruminant. In some cases, the ruminant is bovine, sheep, or goat. In some cases, the source of the casein micelles is from a genetically modified microorganism, for example, yeast, fungi, or bacteria. In some cases, the source of the casein micelles is from a genetically modified plant. In some cases, the genetically modified plant is a soybean plant.

[0016] In some cases, the composition is a dairy product (e.g., milk, cheese), or an intermediate product (e.g., cheese curd) made during the process of making a dairy product. In some cases, casein protein is more than 50% (w/w) of total protein in the composition. In some cases, casein protein is more than 40% (w/w) of total protein in the composition. In some cases, casein protein is more than 30% (w/w) of total protein in the composition. In some cases, casein protein is more than 20% (w/w) of total protein in the composition. In some cases, casein protein is more than 10% (w/w) of total protein in the composition. In some cases, casein protein is more than 5 % (w/w) of total protein in the composition. In some cases, casein protein is less than 50% (w/w) of total protein in the composition. In some cases, casein protein is less than 60% (w/w) of total protein in the composition. In some cases, casein protein is less than 70% (w/w) of total protein in the composition. In some cases, casein protein is less than 80% (w/w) of total protein in the composition. In some cases, casein protein is less than 90% (w/w) of total protein in the composition. In some cases, the weight ratio of total casein protein and total soy protein in the disclosed composition is lower than 0.8. In some cases, the weight ratio of total casein protein and total soy protein is lower than 0.7. In some cases, the weight ratio of total casein protein and total soy protein is lower than 0.6. In some cases, the weight ratio of total casein protein and total soy protein is lower than 0.5. In some cases, the weight ratio of total casein protein and total soy protein is lower than 0.4. In some cases, the weight ratio of total casein protein and total soy protein is lower than 0.3. In some cases, the weight ratio of total casein protein and total soy protein is lower than 0.2. In some cases, the weight ratio of total casein protein and total soy protein is lower than 0.1. In some cases, the weight ratio of total casein protein and total soy protein in the disclosed composition is higher than 0.8. In some cases, the weight ratio of total casein protein and total soy protein is higher than 0.7. In some cases, the weight ratio of total casein protein and total soy protein is higher than 0.6. In some cases, the weight ratio of total casein protein and total soy protein is higher than 0.5. In some cases, the weight ratio of total casein protein and total soy protein is higher than 0.4. In some cases, the weight ratio of total casein protein and total soy protein is higher than 0.3. In some cases, the weight ratio of total casein protein and total soy protein is higher than 0.2. In some cases, the weight ratio of total casein protein and total soy protein is higher than 0.1.

[0017] In some cases, soy protein is more than 30% (w/w) of total protein in the composition. In some cases, soy protein is more than 40% (w/w) of total protein in the composition. In some cases, soy protein is more than 50% (w/w) of total protein in the composition. In some cases, soy protein is more than 60% (w/w) of total protein in the composition. In some cases, soy protein is more than 70% (w/w) of total protein in the composition. In some cases, soy protein is more than 80% (w/w) of total protein in the composition. In some cases, soy protein is more than 90% (w/w) of total protein in the composition. In some cases, the composition is a dairy product or an intermediate product made during the process of making a dairy product (e.g., cheese).

[0018] Some aspects of the disclosure provide a composition comprising a casein micelle and a soy ingredient, wherein the soy ingredient comprises I ls (e.g., glycinin) in an amount greater than 50% (w/w) of total soy proteins in the composition. In some cases, the I ls (e.g., glycinin) is in an amount greater than 60% (w/w) of total soy proteins. In some cases, the I ls (e.g., glycinin) is in an amount greater than 70% (w/w) of total soy proteins. In some cases, the I ls (e.g., glycinin) is in an amount greater than 75% (w/w) of total soy proteins. In some cases, the I ls (e.g., glycinin) is in an amount greater than 80% (w/w) of total soy proteins. In some cases, the I ls (e.g., glycinin) is in an amount greater than 85% (w/w) of total soy proteins. In some cases, the I ls (e.g., glycinin) is in an amount greater than 90% (w/w) of total soy proteins. In some cases, the I ls (e.g., glycinin) is in an amount greater than 95% (w/w) of total soy proteins. In some cases, the I ls (e.g., glycinin) is in an amount greater than 99% (w/w) of total soy proteins. [0019] Some aspects of the disclosure provide a composition comprising a casein micelle and a soy ingredient, wherein the soy ingredient comprises I ls (e.g., glycinin) in an amount greater than 50% (w/w) of total seed storage protein in the composition. In some cases, the I ls (e.g., glycinin) is in an amount greater than 60% (w/w) of total seed storage protein in the composition. In some cases, the I ls (e.g., glycinin) is in an amount greater than 70% (w/w) of total seed storage protein in the composition. In some cases, the I ls (e.g., glycinin) is in an amount greater than 75% (w/w) of total seed storage protein in the composition. In some cases, the I ls (e.g., glycinin) is in an amount greater than 80% (w/w) of total seed storage protein in the composition. In some cases, the I ls (e.g., glycinin) is in an amount greater than 85% (w/w) of total seed storage protein in the composition. In some cases, the I ls (e.g., glycinin) is in an amount greater than 90% (w/w) of total seed storage protein in the composition. In some cases, the I ls (e.g., glycinin) is in an amount greater than 95% (w/w) of total seed storage protein in the composition. In some cases, the I ls (e.g., glycinin) is in an amount greater than 99% (w/w) of total seed storage protein in the composition.

[0020] Some aspects of the disclosure provide a composition comprising a casein micelle and a soy ingredient, wherein the soy ingredient comprises I ls (e.g., glycinin) in an amount greater than 50% (w/w) of total protein in the composition. In some cases, the I ls (e.g., glycinin) is in an amount greater than 60% (w/w) of total protein in the composition. In some cases, the I ls (e.g., glycinin) is in an amount greater than 70% (w/w) of total protein in the composition. In some cases, the I ls (e.g., glycinin) is in an amount greater than 75% (w/w) of total protein in the composition. In some cases, the I ls (e.g., glycinin) is in an amount greater than 80% (w/w) of total protein in the composition. In some cases, the I ls (e.g., glycinin) is in an amount greater than 85% (w/w) of total protein in the composition. In some cases, the I ls (e.g., glycinin) is in an amount greater than 90% (w/w) of total protein in the composition. In some cases, the I ls (e.g., glycinin) is in an amount greater than 95% (w/w) of total protein in the composition. In some cases, the I ls (e.g., glycinin) is in an amount greater than 99% (w/w) of total protein in the composition.

[0021] Some aspects of the disclosure provide a composition comprising a casein micelle and a soy ingredient, wherein the soy ingredient comprises 7s (e.g., 0-conglycinin) in an amount greater than 50% (w/w) of total soy proteins in the composition. In some cases, the 7s (e.g., 0- conglycinin) is in an amount greater than 60% (w/w) of total soy proteins. In some cases, the 7s (e.g., P-conglycinin) is in an amount greater than 70% (w/w) of total soy proteins. In some cases, the 7s (e.g., P-conglycinin) is in an amount greater than 75% (w/w) of total soy proteins. In some cases, the 7s (e.g., P-conglycinin) is in an amount greater than 80% (w/w) of total soy proteins. In some cases, the 7s (e.g., P-conglycinin) is in an amount greater than 85% (w/w) of total soy proteins. In some cases, the 7s (e.g., P-conglycinin) is in an amount greater than 90% (w/w) of total soy proteins. In some cases, the 7s (e.g., P-conglycinin) is in an amount greater than 95% (w/w) of total soy proteins.

[0022] Some aspects of the disclosure provide a composition comprising a casein micelle and a soy ingredient, wherein the soy ingredient comprises 7s (e.g., P-conglycinin) in an amount greater than 50% (w/w) of total protein in the composition. In some cases, the 7s (e.g., 0- conglycinin) is in an amount greater than 60% (w/w) of total protein in the composition. In some cases, the 7s (e.g., P-conglycinin) is in an amount greater than 70% (w/w) of total protein in the composition. In some cases, the 7s (e.g., P-conglycinin) is in an amount greater than 75% (w/w) of total protein in the composition. In some cases, the 7s (e.g., P-conglycinin) is in an amount greater than 80% (w/w) of total protein in the composition. In some cases, the 7s (e.g., 0- conglycinin) is in an amount greater than 85% (w/w) of total protein in the composition. In some cases, the 7s (e.g., P-conglycinin) is in an amount greater than 90% (w/w) of total protein in the composition. In some cases, the 7s (e.g., P-conglycinin) is in an amount greater than 95% (w/w) of total protein in the composition.

[0023] Some aspects of the disclosure provide a composition comprising a casein micelle and a soy ingredient, wherein the soy ingredient comprises 7s (e.g., 0-conglycinin) in an amount greater than 50% (w/w) of total seed storage protein in the composition. In some cases, the 7s (e.g., 0-conglycinin) is in an amount greater than 60% (w/w) of total seed storage protein in the composition. In some cases, the 7s (e.g., P-conglycinin) is in an amount greater than 70% (w/w) of total seed storage protein in the composition. In some cases, the 7s (e.g., P-conglycinin) is in an amount greater than 75% (w/w) of total seed storage protein in the composition. In some cases, the 7s (e.g., P-conglycinin) is in an amount greater than 80% (w/w) of total seed storage protein in the composition. In some cases, the 7s (e.g., P-conglycinin) is in an amount greater than 85% (w/w) of total seed storage protein in the composition. In some cases, the 7s (e.g., P- conglycinin) is in an amount greater than 90% (w/w) of total seed storage protein in the composition. In some cases, the 7s (e.g., P-conglycinin) is in an amount greater than 95% (w/w) of total seed storage protein in the composition.

[0024] Some aspects of the disclosure provide a composition comprising a casein micelle and a soy ingredient, wherein the soy ingredient comprises I ls (e.g., glycinin) in an amount less than 50% (w/w) of total soy proteins in the composition. In some cases, the I ls (e.g., glycinin) is in an amount less than 40% (w/w) of total soy proteins. In some cases, the I ls (e.g., glycinin) is in an amount less than 30% (w/w) of total soy proteins. In some cases, the I ls (e.g., glycinin) is in an amount less than 20% (w/w) of total soy proteins. In some cases, the I ls (e.g., glycinin) is in an amount less than 10% (w/w) of total soy proteins. In some cases, the I ls (e.g., glycinin) is in an amount less than 5% (w/w) of total soy proteins. In some cases, the I ls (e.g., glycinin) is in an amount less than 1% (w/w) of total soy proteins. In some cases, the I ls (e.g., glycinin) is in an amount less than 0.1% (w/w) of total soy proteins. In some cases, the I ls (e.g., glycinin) is in an amount less than 0.01% (w/w) of total soy proteins. In some cases, the I ls (e.g., glycinin) is in an amount less than 0.001% (w/w) of total soy proteins. In some cases, the I ls (e.g., glycinin) is in an amount less than 0.0001% (w/w) of total soy proteins. In some cases, the I ls (e.g., glycinin) is in an amount less than 0.00001% (w/w) of total soy proteins. In some cases, the I ls (e.g., glycinin) is in an amount less than 0.000001% (w/w) of total soy proteins.

[0025] Some aspects of the disclosure provide a composition comprising a casein micelle and a soy ingredient, wherein the soy ingredient comprises I ls (e.g., glycinin) in an amount less than 50% (w/w) of total protein in the composition. In some cases, the I ls (e.g., glycinin) is in an amount less than 40% (w/w) of total protein in the composition. In some cases, the I ls (e.g., glycinin) is in an amount less than 30% (w/w) of total protein in the composition. In some cases, the I ls (e.g., glycinin) is in an amount less than 20% (w/w) of total protein in the composition. In some cases, the I ls (e.g., glycinin) is in an amount less than 10% (w/w) of total protein in the composition. In some cases, the I ls (e.g., glycinin) is in an amount less than 5% (w/w) of total protein in the composition. In some cases, the I ls (e.g., glycinin) is in an amount less than 1% (w/w) of total protein in the composition. In some cases, the I ls (e.g., glycinin) is in an amount less than 0.1% (w/w) of total protein in the composition. In some cases, the I ls (e.g., glycinin) is in an amount less than 0.01% (w/w) of total protein in the composition. In some cases, the I ls (e.g., glycinin) is in an amount less than 0.001% (w/w) of total protein in the composition. In some cases, the I ls (e.g., glycinin) is in an amount less than 0.0001% (w/w) of total protein in the composition. In some cases, the I ls (e.g., glycinin) is in an amount less than 0.00001% (w/w) of total protein in the composition. In some cases, the I ls (e.g., glycinin) is in an amount less than 0.000001% (w/w) of total protein in the composition.

[0026] Some aspects of the disclosure provide a composition comprising a casein micelle and a soy ingredient, wherein the soy ingredient comprises I ls (e.g., glycinin) in an amount less than 50% (w/w) of total seed storage protein in the composition. In some cases, the I ls (e.g., glycinin) is in an amount less than 40% (w/w) of total seed storage protein in the composition. In some cases, the I ls (e.g., glycinin) is in an amount less than 30% (w/w) of total seed storage protein in the composition. In some cases, the I ls (e.g., glycinin) is in an amount less than 20% (w/w) of total seed storage protein in the composition. In some cases, the I ls (e.g., glycinin) is in an amount less than 10% (w/w) of total seed storage protein in the composition. In some cases, the I ls (e.g., glycinin) is in an amount less than 5% (w/w) of total seed storage protein in the composition. In some cases, the I ls (e.g., glycinin) is in an amount less than 1% (w/w) of total seed storage protein in the composition. In some cases, the I ls (e.g., glycinin) is in an amount less than 0.1% (w/w) of total seed storage protein in the composition. In some cases, the I ls (e.g., glycinin) is in an amount less than 0.01% (w/w) of total seed storage protein in the composition. In some cases, the I ls (e.g., glycinin) is in an amount less than 0.001% (w/w) of total seed storage protein in the composition. In some cases, the I ls (e.g., glycinin) is in an amount less than 0.0001% (w/w) of total seed storage protein in the composition. In some cases, the I ls (e.g., glycinin) is in an amount less than 0.00001% (w/w) of total seed storage protein in the composition. In some cases, the I ls (e.g., glycinin) is in an amount less than 0.000001% (w/w) of total seed storage protein in the composition.

[0027] Some aspects of the disclosure provide a composition comprising a casein micelle and a soy ingredient, wherein the soy ingredient comprises 7s (e.g., 0-conglycinin) in an amount less than 50% (w/w) of total soy proteins in the composition. In some cases, the 7s (e.g., 0- conglycinin) is in an amount less than 40% (w/w) of total soy proteins. In some cases, the 7s (e.g., 0-conglycinin) is in an amount less than 30% (w/w) of total soy proteins. In some cases, the 7s (e.g., 0-conglycinin) is in an amount less than 20% (w/w) of total soy proteins. In some cases, the 7s (e.g., 0-conglycinin) is in an amount less than 10% (w/w) of total soy proteins. In some cases, the 7s (e.g., 0-conglycinin) is in an amount less than 5% (w/w) of total soy proteins. In some cases, the 7s (e.g., 0-conglycinin) is in an amount less than 1% (w/w) of total soy proteins. In some cases, the 7s (e.g., 0-conglycinin) is in an amount less than 0.1% (w/w) of total soy proteins. In some cases, the 7s (e.g., P-conglycinin) is in an amount less than 0.01% (w/w) of total soy proteins. In some cases, the 7s (e.g., P-conglycinin) is in an amount less than 0.001% (w/w) of total soy proteins. In some cases, the 7s (e.g., P-conglycinin) is in an amount less than 0.0001% (w/w) of total soy proteins. In some cases, the 7s (e.g., P-conglycinin) is in an amount less than 0.00001% (w/w) of total soy proteins. In some cases, the 7s (e.g., P-conglycinin) is in an amount less than 0.000001% (w/w) of total soy proteins.

[0028] Some aspects of the disclosure provide a composition comprising a casein micelle and a soy ingredient, wherein the soy ingredient comprises 7s (e.g., P-conglycinin) in an amount less than 50% (w/w) of total protein in the composition. In some cases, the 7s (e.g., P-conglycinin) is in an amount less than 40% (w/w) of total protein in the composition. In some cases, the 7s (e.g., P-conglycinin) is in an amount less than 30% (w/w) of total protein in the composition. In some cases, the 7s (e.g., P-conglycinin) is in an amount less than 20% (w/w) of total protein in the composition. In some cases, the 7s (e.g., P-conglycinin) is in an amount less than 10% (w/w) of total protein in the composition. In some cases, the 7s (e.g., P-conglycinin) is in an amount less than 5% (w/w) of total protein in the composition. In some cases, the 7s (e.g., P-conglycinin) is in an amount less than 1% (w/w) of total protein in the composition. In some cases, the 7s (e.g., P-conglycinin) is in an amount less than 0.1% (w/w) of total protein in the composition. In some cases, the 7s (e.g., P-conglycinin) is in an amount less than 0.01% (w/w) of total protein in the composition. In some cases, the 7s (e.g., P-conglycinin) is in an amount less than 0.001% (w/w) of total protein in the composition. In some cases, the 7s (e.g., P-conglycinin) is in an amount less than 0.0001% (w/w) of total protein in the composition. In some cases, the 7s (e.g., P- conglycinin) is in an amount less than 0.00001% (w/w) of total protein in the composition. In some cases, the 7s (e.g., P-conglycinin) is in an amount less than 0.000001% (w/w) of total protein in the composition.

[0029] Some aspects of the disclosure provide a composition comprising a casein micelle and a soy ingredient, wherein the soy ingredient comprises 7s (e.g., P-conglycinin) in an amount less than 50% (w/w) of total seed storage protein in the composition. In some cases, the 7s (e.g., P- conglycinin) is in an amount less than 40% (w/w) of total seed storage protein in the composition. In some cases, the 7s (e.g., P-conglycinin) is in an amount less than 30% (w/w) of total seed storage protein in the composition. In some cases, the 7s (e.g., P-conglycinin) is in an amount less than 20% (w/w) of total seed storage protein in the composition. In some cases, the 7s (e.g., P-conglycinin) is in an amount less than 10% (w/w) of total seed storage protein in the composition. In some cases, the 7s (e.g., P-conglycinin) is in an amount less than 5% (w/w) of total seed storage protein in the composition. In some cases, the 7s (e.g., P-conglycinin) is in an amount less than 1% (w/w) of total seed storage protein in the composition. In some cases, the 7s (e.g., P-conglycinin) is in an amount less than 0.1% (w/w) of total seed storage protein in the composition. In some cases, the 7s (e.g., P-conglycinin) is in an amount less than 0.01% (w/w) of total seed storage protein in the composition. In some cases, the 7s (e.g., P-conglycinin) is in an amount less than 0.001% (w/w) of total seed storage protein in the composition. In some cases, the 7s (e.g., P-conglycinin) is in an amount less than 0.0001% (w/w) of total seed storage protein in the composition. In some cases, the 7s (e.g., P-conglycinin) is in an amount less than 0.00001% (w/w) of total seed storage protein in the composition. In some cases, the 7s (e.g., P- conglycinin) is in an amount less than 0.000001% (w/w) of total seed storage protein in the composition.

[0030] In some cases, the total soy protein comprises one or more of seed storage proteins, for example, legumin, vicilin, prolamin, gliadin, P-conglycinin, or glycinin. In some cases, the composition is a dairy product (e.g., cheese) or an intermediate product made during the process of making a dairy product.

[0031] In some cases, the composition forms a cheese curd in a cheese making process. In some cases, the composition forms a firm cheese curd in a cheese making process. In some cases, the composition forms a firm cheese curd in a cheese making process within 15 minutes in the cheese making process. In some cases, the composition forms a firm cheese curd in a cheese making process within 30 minutes in the cheese making process. In some cases, the composition forms a firm cheese curd in a cheese making process within 60 minutes in the cheese making process. In some cases, the composition forms a firm cheese curd in a cheese making process within 100 minutes in the cheese making process.

[0032] In some cases, the composition forms a soft cheese curd in a cheese making process. In some cases, the composition forms a soft cheese curd within 15 minutes in the cheese making process. In some cases, the composition forms a soft cheese curd within 30 minutes in the cheese making process. In some cases, the composition forms a soft cheese curd within 60 minutes in the cheese making process. In some cases, the composition forms a soft cheese curd within 100 minutes in the cheese making process.

[0033] In some cases, the cheese making process is performed at pH between 6.0-7.0. In some cases, the cheese making process is performed at pH between 6.0-6.5. In some cases, the cheese making process is performed at pH between 6.5-7.0. In some cases, the cheese making process comprises adding an enzyme (e.g., one or more enzymes found in rennet, or other suitable proteases) to the composition to cause the casein micelle to precipitate.

[0034] In some aspects, the current disclosure provides a cheese product comprising the composition disclosed herein. In some aspects, the current disclosure provides a method of making cheese, comprising providing the composition disclosed herein, and adding rennet to the composition to cause the casein micelles to precipitate. In some aspects, the method disclosed herein comprises mixing soy ingredient with soluble casein micelles.

[0035] In some aspects, the current disclosure provides compositions, methods and systems for making cheese using a modified soy ingredient. In some aspects, the modified soy ingredient comprises a non-detectable or reduced level of an anti -nutritional or toxin in soybean (for example, including trypsin inhibitors, phytic acid, lectins, or soybean toxin), compared to a natural occurring level found in non-modified soybeans. For example, the non-detectable or reduced levels of anti -nutritional or toxin can be achieved by knocking out or down one or more of the following genes: anti-nutritional/toxins in soybean, including trypsin inhibitors, phytic acid, lectins, soybean toxin.

[0036] In some aspects, the current disclosure provides compositions, methods and systems for making cheese using one or more seed storage proteins (e.g., legumin, vicilin, prolamin gliadin, etc.), in lieu of soy proteins as disclosed herein.

[0037] In some aspects, the current disclosure provides a method of making a dairy product, comprising, providing a liquid mixture comprising casein micelles and at least one soy protein; removing a portion of the soy protein from the liquid mixture; and adding an enzyme to the liquid mixture to cause the casein micelles to precipitate. In some cases, the at least one soy protein to be removed comprises a subunit of conglycinin. In some cases, removing the portion of the soy protein from the liquid mixture decreases the ratio of conglycinin to glycinin (conglycinin/glycinin). In some cases, the enzyme comprises at least one enzyme found in rennet. In some cases, the enzyme comprises a protease. In some cases, the enzyme comprises at least one of chymosin, pepsin or lipase.

[0038] In some cases, removing the portion of soy protein from the mixture comprises adding a salt to the liquid mixture to precipitate the portion of soy protein. In some cases, the salt is at least one of sodium phosphate, calcium chloride, or potassium chloride. In some cases, removing the portion of soy protein from the mixture further comprises filtering the composition to produce a supernatant.

[0039] In some cases, filtering the composition comprises using a microfiltration (MF) membrane to filter the composition. In some cases, filtering the composition comprises using a benchtop tangential flow filtration system to filter the composition. In some cases, the microfiltration (MF) membrane has a pore size between 0.1 and 10 pm. In some cases, the microfiltration (MF) membrane has a pore size between 0.1 and 1 pm. In some cases, microfiltration (MF) membrane has a pore size between 1 and 10 pm. In some cases, the casein micelles precipitate to form a curd. In some cases, the casein micelles precipitate to form a solid or semi-solid. [0040] Some aspects of the disclosure provide a composition, comprising a casein micelle derived from a plant; and a plant protein comprising one of legumin, vicilin, prolamin, gliadin, 0- conglycinin, or glycinin, or any combination thereof. In some cases, the plant protein is a seed storage protein. In some cases, one or more of the plant proteins described herein is a soy protein. In some cases, the plant protein described herein is non-soy protein, including, for example, a protein from maize, pea, peanut, rice, sesame, lima bean, pea, chickpea, maize, wheat, rice, barley, or oat, etc. In some cases, the plant protein comprises I ls (e.g., glycinin). In some cases, the plant protein further comprises 7s (e.g., 0-conglycinin). In some cases, the ratio of 7s/l Is is lower than a naturally occurring ratio. In some cases, the ratio of 7s/l Is (w/w) is lower than 0.5. In some cases, the ratio of 7s/l Is (w/w) is lower than 0.75. In some cases, the ratio of 7s/l Is (w/w) is lower than 1. In some cases, the ratio of 7s/l Is (w/w) is lower than 1.3. In some cases, the ratio of 7s/l 1 s(w/w) is lower than 0.4. In some cases, the ratio of 7s/l Is (w/w) is lower than 0.3. In some cases, the ratio of 7s/l Is (w/w) is lower than 0.25. In some cases, the ratio of 7s/l Is (w/w) is lower than 0.2. In some cases, the ratio of 7s/l Is (w/w) is lower than 0.1. In some cases, the ratio of 7s/l Is (w/w) is lower than 0.05. In some cases, the ratio of 7s/l Is (w/w) is lower than 0.01. In some cases, the ratio of 7s/l Is (w/w) is lower than 00.1. In some cases, the composition does not comprise a detectable amount of 7s (e.g., 0-conglycinin).

[0041] In some cases, the ratio of 11 s/7s is lower than a naturally occurring ratio. In some cases, the ratio of 11 s/7s (w/w) is lower than 0.5. In some cases, the ratio of 11 s/7s (w/w) is lower than 0.75. In some cases, the ratio of 11 s/7s (w/w) is lower than 1. In some cases, the ratio of 11 s/7s (w/w) is lower than 1.3. In some cases, the ratio of 7s/l 1 s(w/w) is lower than 0.4. In some cases, the ratio of 11 s/7s (w/w) is lower than 0.3. In some cases, the ratio of 11 s/7s (w/w) is lower than 0.25. In some cases, the ratio of 11 s/7s (w/w) is lower than 0.2. In some cases, the ratio of 11 s/7s (w/w) is lower than 0.1. In some cases, the ratio of 1 ls/7s (w/w) is lower than 0.01. In some cases, the ratio of 11 s/7s (w/w) is lower than 0.001. In some cases, the ratio of 11 s/7s (w/w) is lower than 0.0001. In some cases, the ratio of 11 s/7s (w/w) is lower than 0.00001. In some cases, the ratio of 11 s/7s (w/w) is lower than 0.000001. In some cases, the ratio of 11 s/7s (w/w) is lower than 0.0000001. In some cases, the composition does not comprise a detectable amount of 1 Is (e.g., glycinin).

[0042] In some instances, the plant protein is inside the casein micelle. In some cases, an outer layer of the casein micelle is enriched with a K-casein, and an inner matrix of the casein micelle comprises at least one of aS 1 -casein, aS2-casein, or 0-casein. In some cases, the plant protein (e.g., 7s, I ls, or both) interacts with one or more of the casein proteins in the inner matrix. In some cases, the plant protein binds to any one of aS 1 -casein, aS2-casein, or 0-casein. In some instances, the binding between the plant protein and the aS 1 -casein, aS2-casein, or 0-casein is noncovalent. In some cases, the plant protein is at least partially embedded in the inner matrix. In some cases, the plant protein is fully embedded in the inner matrix. In some cases, the plant protein is on the surface of the casein micelle. In some cases, the plant protein (e.g., 7s, 1 Is, or both) interacts with a K-casein on an outer layer of the casein micelle. In some cases, the plant protein binds to the K-casein. In some instances, the binding between the plant protein and the K- casein is noncovalent. In some cases, the plant protein is at least partially embedded in the outer layer of the casein micelle. In some cases, the composition is a dairy product or an intermediate product made during the process of making a dairy product (e.g., cheese).

[0043] In some cases, the plant protein that is present inside the casein micelle or attached to the surface of the casein micelle is present in a trace amount in the composition, as described herein. For example, in some cases, the trace amount plant protein is less than 5% (w/w) of total protein in the composition. In some cases, the trace amount plant protein is less than 4% (w/w) of total protein in the composition. In some cases, the trace amount plant protein is less than 3% (w/w) of total protein in the composition. In some cases, the trace amount plant protein is less than 2% (w/w) of total protein in the composition. In some cases, the trace amount plant protein is less than 1% (w/w) of total protein in the composition. In some cases, the trace amount plant protein is less than 0.5% (w/w) of total protein in the composition. In some cases, the trace amount plant protein is less than 0.4% (w/w) of total protein in the composition. In some cases, the trace amount plant protein is less than 0.3% (w/w) of total protein in the composition. In some cases, the trace amount plant protein is less than 0.2% (w/w) of total protein in the composition. In some cases, the trace amount plant protein is less than 0.1% (w/w) of total protein in the composition. In some cases, the trace amount plant protein is less than 0.09% (w/w) of total protein in the composition. In some cases, the trace amount plant protein is less than 0.08% (w/w) of total protein in the composition. In some cases, the trace amount plant protein is less than 0.07% (w/w) of total protein in the composition. In some cases, the trace amount plant protein is less than 0.06% (w/w) of total protein in the composition. In some cases, the trace amount plant protein is less than 0.05% (w/w) of total protein in the composition. In some cases, the trace amount plant protein is less than 0.04% (w/w) of total protein in the composition. In some cases, the trace amount plant protein is less than 0.03% (w/w) of total protein in the composition. In some cases, the trace amount plant protein is less than 0.02% (w/w) of total protein in the composition. In some cases, the trace amount plant protein is less than 0.01% (w/w) of total protein in the composition. In some cases, the trace amount plant protein is less than 0.005% (w/w) of total protein in the composition. In some cases, the trace amount plant protein is less than 0.001% (w/w) of total protein in the composition. In some cases, the trace amount plant protein is less than 0.0005% (w/w) of total protein in the composition. In some cases, the trace amount plant protein is less than 0.0001% (w/w) of total protein in the composition. In some cases, the trace amount plant protein is less than 0.00005% (w/w) of total protein in the composition. In some cases, the trace amount plant protein is more than 4% (w/w) of total protein in the composition. In some cases, the trace amount plant protein is more than 3% (w/w) of total protein in the composition. In some cases, the trace amount plant protein is more than 2% (w/w) of total protein in the composition. In some cases, the trace amount plant protein is more than 1% (w/w) of total protein in the composition. In some cases, the trace amount plant protein is more than 0.5% (w/w) of total protein in the composition. In some cases, the trace amount plant protein is more than 0.4% (w/w) of total protein in the composition. In some cases, the trace amount plant protein is more than 0.3% (w/w) of total protein in the composition. In some cases, the trace amount plant protein is more than 0.2% (w/w) of total protein in the composition. In some cases, the trace amount plant protein is more than 0.1% (w/w) of total protein in the composition. In some cases, the trace amount plant protein is more than 0.09% (w/w) of total protein in the composition. In some cases, the trace amount plant protein is more than 0.08% (w/w) of total protein in the composition. In some cases, the trace amount plant protein is more than 0.07% (w/w) of total protein in the composition. In some cases, the trace amount plant protein is more than 0.06% (w/w) of total protein in the composition. In some cases, the trace amount plant protein is more than 0.05% (w/w) of total protein in the composition. In some cases, the trace amount plant protein is more than 0.04% (w/w) of total protein in the composition. In some cases, the trace amount plant protein is more than 0.03% (w/w) of total protein in the composition. In some cases, the trace amount plant protein is more than 0.02% (w/w) of total protein in the composition. In some cases, the trace amount plant protein is more than 0.01% (w/w) of total protein in the composition. In some cases, the trace amount plant protein is more than 0.005% (w/w) of total protein in the composition. In some cases, the trace amount plant protein is more than 0.001% (w/w) of total protein in the composition. In some cases, the trace amount plant protein more than 0.0005% (w/w) of total protein in the composition. In some cases, the trace amount plant protein is more than 0.0001% (w/w) of total protein in the composition. In some cases, the trace amount plant protein is more than 0.00005% (w/w) of total protein in the composition. In some cases, the trace amount plant protein is more than 0.00003% (w/w) of total protein in the composition. In some cases, the trace amount plant protein is more than 0.00001% (w/w) of total protein in the composition. In some cases, the composition is a dairy product or an intermediate product made during the process of making a dairy product (e.g., cheese).

[0044] In some cases, the plant protein present in the compositions disclosed herein is not in a trace amount. For example, in some cases, the plant protein is more than 5% (w/w) of total protein content in the composition. In some cases, the plant protein is more than 10% (w/w) of total protein content in the composition. In some cases, the plant protein is more than 20% (w/w) of total protein content in the composition. In some cases, the plant protein is more than 30% (w/w) of total protein content in the composition. In some cases, the plant protein is more than 40% (w/w) of total protein content in the composition. In some cases, the plant protein is more than 50% (w/w) of total protein content in the composition. In some cases, the plant protein is more than 60% (w/w) of total protein content in the composition. In some cases, the plant protein is more than 70% (w/w) of total protein content in the composition. In some cases, the plant protein is more than 80% (w/w) of total protein content in the composition. In some cases, the plant protein is less than 10% (w/w) of total protein content in the composition. In some cases, the plant protein is less than 20% (w/w) of total protein content in the composition. In some cases, the plant protein is less than 30% (w/w) of total protein content in the composition. In some cases, the plant protein is less than 40% (w/w) of total protein content in the composition. In some cases, the plant protein is less than 50% (w/w) of total protein content in the composition. In some cases, the plant protein is less than 60% (w/w) of total protein content in the composition. In some cases, the plant protein is less than 70% (w/w) of total protein content in the composition. In some cases, the plant protein is less than 80% (w/w) of total protein content in the composition. In some cases, the plant protein is less than 90% (w/w) of total protein content in the composition. In some cases, the composition is a dairy product or an intermediate product made during the process of making a dairy product (e.g., cheese).

[0045] In some cases, the plant protein that is present inside the casein micelle or attached to the surface of the casein micelle is present in a trace amount in the composition, as described herein. For example, in some cases, the trace amount plant protein is less than 5% (w/w) of casein protein weight in the composition. In some cases, the trace amount plant protein is less than 4% (w/w) of casein protein weight in the composition. In some cases, the trace amount plant protein is less than 3% (w/w) of casein protein weight in the composition. In some cases, the trace amount plant protein is less than 2% (w/w) of casein protein weight in the composition. In some cases, the trace amount plant protein is less than 1% (w/w) of casein protein weight in the composition. In some cases, the trace amount plant protein is less than 0.5% (w/w) of casein protein weight in the composition. In some cases, the trace amount plant protein is less than 0.4% (w/w) of casein protein weight in the composition. In some cases, the trace amount plant protein is less than 0.3% (w/w) of casein protein weight in the composition. In some cases, the trace amount plant protein is less than 0.2% (w/w) of casein protein weight in the composition. In some cases, the trace amount plant protein is less than 0.1% (w/w) of casein protein weight in the composition. In some cases, the trace amount plant protein is less than 0.09% (w/w) of casein protein weight in the composition. In some cases, the trace amount plant protein is less than 0.08% (w/w) of casein protein weight in the composition. In some cases, the trace amount plant protein is less than 0.07% (w/w) of casein protein weight in the composition. In some cases, the trace amount plant protein is less than 0.06% (w/w) of casein protein weight in the composition. In some cases, the trace amount plant protein is less than 0.05% (w/w) of casein protein weight in the composition. In some cases, the trace amount plant protein is less than 0.04% (w/w) of casein protein weight in the composition. In some cases, the trace amount plant protein is less than 0.03% (w/w) of casein protein weight in the composition. In some cases, the trace amount plant protein is less than 0.02% (w/w) of casein protein weight in the composition. In some cases, the trace amount plant protein is less than 0.01% (w/w) of casein protein weight in the composition. In some cases, the trace amount plant protein is less than 0.005% (w/w) of casein protein weight in the composition. In some cases, the trace amount plant protein is less than 0.001% (w/w) of casein protein weight in the composition. In some cases, the trace amount plant protein is less than 0.0005% (w/w) of casein protein weight in the composition. In some cases, the trace amount plant protein is less than 0.0001% (w/w) of casein protein weight in the composition. In some cases, the trace amount plant protein is less than 0.00005% (w/w) of casein protein weight in the composition. In some cases, the trace amount plant protein is more than 4% (w/w) of casein protein weight in the composition. In some cases, the trace amount plant protein is more than 3% (w/w) of casein protein weight in the composition. In some cases, the trace amount plant protein is more than 2% (w/w) of casein protein weight in the composition. In some cases, the trace amount plant protein is more than 1% (w/w) of casein protein weight in the composition. In some cases, the trace amount plant protein is more than 0.5% (w/w) of casein protein weight in the composition. In some cases, the trace amount plant protein is more than 0.4% (w/w) of casein protein weight in the composition. In some cases, the trace amount plant protein is more than 0.3% (w/w) of casein protein weight in the composition. In some cases, the trace amount plant protein is more than 0.2% (w/w) of casein protein weight in the composition. In some cases, the trace amount plant protein is more than 0.1% (w/w) of casein protein weight in the composition. In some cases, the trace amount plant protein is more than 0.09% (w/w) of casein protein weight in the composition. In some cases, the trace amount plant protein is more than 0.08% (w/w) of casein protein weight in the composition. In some cases, the trace amount plant protein is more than 0.07% (w/w) of casein protein weight in the composition. In some cases, the trace amount plant protein is more than 0.06% (w/w) of casein protein weight in the composition. In some cases, the trace amount plant protein is more than 0.05% (w/w) of casein protein weight in the composition. In some cases, the trace amount plant protein is more than 0.04% (w/w) of casein protein weight in the composition. In some cases, the trace amount plant protein is more than 0.03% (w/w) of casein protein weight in the composition. In some cases, the trace amount plant protein is more than 0.02% (w/w) of casein protein weight in the composition. In some cases, the trace amount plant protein is more than 0.01% (w/w) of casein protein weight in the composition. In some cases, the trace amount plant protein is more than 0.005% (w/w) of casein protein weight in the composition. In some cases, the trace amount plant protein is more than 0.001% (w/w) of casein protein weight in the composition. In some cases, the trace amount plant protein more than 0.0005% (w/w) of casein protein weight in the composition. In some cases, the trace amount plant protein is more than 0.0001% (w/w) of casein protein weight in the composition. In some cases, the trace amount plant protein is more than 0.00005% (w/w) of casein protein weight in the composition. In some cases, the trace amount plant protein is more than 0.00003% (w/w) of casein protein weight in the composition. In some cases, the trace amount plant protein is more than 0.00001% (w/w) of casein protein weight in the composition. In some cases, the composition is a dairy product or an intermediate product made during the process of making a dairy product (e.g., cheese).

[0046] In some cases, the plant protein present in the compositions disclosed herein (e.g., a dairy product, or an intermediate product made during the process of making a dairy product (e.g., cheese)) is not in a trace amount. For example, in some cases, the plant protein is more than 5% (w/w) of casein protein weight in the composition. In some cases, the plant protein is more than 10% (w/w) of casein protein weight in the composition. In some cases, the plant protein is more than 20% (w/w) of casein protein weight in the composition. In some cases, the plant protein is more than 30% (w/w) of casein protein weight in the composition. In some cases, the plant protein is more than 40% (w/w) of casein protein weight in the composition. In some cases, the plant protein is more than 50% (w/w) of casein protein weight in the composition. In some cases, the plant protein is more than 60% (w/w) of casein protein weight in the composition. In some cases, the plant protein is more than 70% (w/w) of casein protein weight in the composition. In some cases, the plant protein is more than 80% (w/w) of casein protein weight in the composition. In some cases, the plant protein is less than 10% (w/w) of casein protein weight in the composition. In some cases, the plant protein is less than 20% (w/w) of casein protein weight in the composition. In some cases, the plant protein is less than 30% (w/w) of casein protein weight in the composition. In some cases, the plant protein is less than 40% (w/w) of casein protein weight in the composition. In some cases, the plant protein is less than 50% (w/w) of casein protein weight in the composition. In some cases, the plant protein is less than 60% (w/w) of casein protein weight in the composition. In some cases, the plant protein is less than 70% (w/w) of casein protein weight in the composition. In some cases, the plant protein is less than 80% (w/w) of casein protein weight in the composition. In some cases, the plant protein is less than 90% (w/w) of casein protein weight in the composition. In some cases, the composition is a dairy product or an intermediate product made during the process of making a dairy product (e.g., cheese).

[0047] In some cases, the number of plant proteins inside the casein micelle over the number of casein proteins of the casein micelle is less than 1 :50000, less than 1 : 40000, less than 1 : 30000, less than 1 : 20000, less than 1 : 10000, less than 1 : 9000, less than 1 : 8000, less than 1 : 7000, less than 1 : 6000, less than 1 : 5000, less than 1 : 4000, less than 1 : 3000, less than 1 : 2000, less than 1 : 1000, less than 1 : 900, less than 1 : 800, less than 1 : 700, less than 1 : 600, less than 1 : 500, less than 1 : 400, less than 1 : 300, less than 1 : 200, less than 1 : 100, less than 1 : 50, less than 1 : 10, less than 1 : 9, less than 1 : 8, less than 1 : 7, less than 1 : 6, less than 1 : 5, less than 1 : 4, less than 1 : 3, less than 1 : 2, or less than 1. In some cases, the number of plant proteins inside the casein micelle over the number of casein proteins of the casein micelle is more than 1 : 50000, more than 1 : 40000, more than 1 : 30000, more than 1 : 20000, more than 1 : 10000, more than 1 : 9000, more than 1 : 8000, more than 1 : 7000, more than 1 : 6000, more than 1 : 5000, more than 1 : 4000, more than 1 : 3000, more than 1 : 2000, more than 1 : 1000, more than 1 : 900, more than 1 : 800, more than 1 : 700, more than 1 : 600, more than 1 : 500, more than 1 : 400, more than 1 : 300, more than 1 : 200, more than 1 : 100, more than 1 : 50, more than 1 : 10, more than 1 : 9, more than 1 : 8, more than 1 : 7, more than 1 : 6, more than 1 : 5, more than 1 : 4, more than 1 : 3, more than 1 : 2, or more than 1.

[0048] In some cases, the composition does not comprise a detectable amount of a-lactalbumin. [0049] In some cases, the composition does not comprise a detectable amount of P-lactoglobulin. In some cases, the composition does not comprise a detectable amount of a-S2-casein. In some cases, the composition does not comprise a detectable amount of lactoferrin. In some cases, the composition does not comprise a detectable amount of transferrin. In some cases, the composition does not comprise a detectable amount of serum albumin. In some cases, the composition does not comprise a detectable amount of lysozyme. In some cases, the composition does not comprise a detectable amount of lactoperoxidase. In some cases, the composition does not comprise a detectable amount of immunoglobulin-A. In some cases, the composition does not comprise a detectable amount of lipase.

[0050] In some cases, the composition is free of a-lactalbumin. In some cases, the composition is free of P-lactoglobulin. In some cases, the composition is free of a-S2-casein. In some cases, the composition is free of a-Sl -casein. In some cases, the composition is free of P-casein. In some cases, the composition is free of lactoferrin. In some cases, the composition is free of transferrin. In some cases, the composition is free of serum albumin. In some cases, the composition is free of lysozyme. In some cases, the composition is free of lactoperoxidase. In some cases, the composition is free of immunoglobulin-A. In some cases, the composition is free of lipase. [0051] In some cases, the composition is essentially free of a-lactalbumin. In some cases, the composition is essentially free of P-lactoglobulin. In some cases, the composition is essentially free of a-S2-casein. In some cases, the composition is essentially free of P-casein. In some cases, the composition is essentially free of a-Sl -casein. In some cases, the composition is essentially free of lactoferrin. In some cases, the composition is essentially free of transferrin. In some cases, the composition is essentially free of serum albumin. In some cases, the composition is essentially free of lysozyme. In some cases, the composition is essentially free of lactoperoxidase. In some cases, the composition is essentially free of immunoglobulin-A. In some cases, the composition is essentially free of lipase.

[0052] In some cases, the composition further comprises a coagulant. In some cases, the coagulant is in a detectable amount. In some cases, the coagulant is a magnesium salt. In some cases, the magnesium salt is at least one of magnesium chloride (MgC12) or magnesium sulfate (MgSO4). In some cases, the coagulant is a calcium salt. In some cases, the calcium salt is at least one of calcium chloride (CaC12) or calcium sulfate (CaSO4). In some cases, the coagulant is glucono-delta-1 actone (GDL). In some cases, the coagulant is less than 0.1% (w/w) the composition. In some cases, the coagulant is less than 0.05% (w/w) the composition. In the coagulant is less than 0.01% (w/w) the composition. In some cases, the coagulant is less than 0.005% (w/w) the composition. In some cases, the coagulant is less than 0.001% (w/w) the composition. In some cases, the coagulant is more than 0.0005% (w/w) the composition. In some cases, the coagulant is more than 0.0001% (w/w) the composition. In some cases, the composition further comprises a fat. In some cases, the composition has a total fat content below 1%. In some cases, the fat is plant fat. In some cases, the composition has a total fat content below 0.1%. In some cases, the composition has a total fat content below 0.01%. In some cases, the composition does not comprise a detectable amount of animal fat. In some cases, the composition does not comprise a detectable amount of lactose. In some cases, the composition does not comprise a detectable amount of animal hormone. In some cases, the composition does not comprise a detectable amount of estrogen. In some cases, the composition does not comprise a detectable amount of progesterone. In some cases, the composition does not comprise a detectable amount of corticoid. In some cases, the composition does not comprise a detectable amount of androgen. In some cases, the composition is a substitute dairy product. In some instances, the composition is a powder. In some cases, the casein micelles derived from plants confer on the composition one or more characteristics of a dairy product selected from the group consisting of: taste, flavor, aroma, appearance, mouthfeel, density, structure, texture, elasticity, springiness, coagulation, binding, leavening, aeration, foaming, creaminess, and emulsification. In some cases, the dairy product is at least one of milk, cheese, yogurt, ghee, or butter.

[0053] In some instances, this disclosure provides methods of producing curds with improved elasticity over curds made with naturally-occurring ratios of soy proteins. In some cases, a ratio of conglycinin to glycinin provided herein produces curds with an elasticity of greater than 60%, 70%, 80%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 99.5%, or 100%. In some cases, the ratio of conglycinin to glycinin is greater than 60%, 70%, 80%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, or 99.5%, or 100%, combined with a composition with a percentage of casein that is at least 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90% , 95%, 96%, 97%, 98%, 99%, or 99.5%, or 100% of total protein within the composition. For example, in some cases, the composition may have about 95% casein and about 5% total soy protein, or about 85% casein and about 15% total soy protein.

[0054] In some instances, the composition further comprises a solvent. In some cases, the solvent is water. In some case, the composition further comprises a plant lipid, a plant sugar, a plant polyphenol, or plant DNA molecule, or any combination thereof. In some case, the plant lipid, a plant sugar, a plant polyphenol, or plant DNA molecule is present in a trace amount in the composition. In some cases, the plant lipoid comprises at least one of stigmasterol, sitosterol, campesterol, or brassicasterol. In some cases, the plant polyphenol comprises isoflavone. In some cases, the plant DNA comprises plant DNA fragments.

[0055] The current disclosure provides compositions, methods, and systems for separating recombinant protein expressed in a plant from other plant materials.

[0056] In some aspects, the current disclosure provides methods for extracting one or more recombinant proteins expressed in a plant, comprising obtaining a mixture comprising water, the recombinant protein, and a component from the plant; adding a coagulant to the mixture to cause at least a portion of the plant component to coagulant; and extracting supernatant from the mixture, wherein the supernatant comprises the recombinant protein. In some aspects, the current disclosure provides methods for extracting a recombinant casein micelle expressed in a plant, comprising obtaining a mixture comprising water, the recombinant protein, and a component from the plant; adding a coagulant to the mixture to cause at least a portion of the plant component to coagulant; and extracting supernatant from the mixture, wherein the supernatant comprises the recombinant protein. Unexpectedly, under the conditions disclosed herein, only plant proteins coagulated, while the casein protein(s) and/or micelles remain soluble. In some cases, the methods are performed in sequence. In some cases, the mixture comprises a slurry made by blending the plant in water. In some cases, the disclosed methods further comprise separating insoluble plant components from the slurry to produce a plant-based milk before adding the coagulant to the mixture.

[0057] In some cases, the plant is a legume. In some cases, the plant is at least one of alfalfa, cassava, cotton, cowpea, maize, pea, peanut, rice, sesame, sorghum, soybean, or yam. In some cases, the plant component comprises a plant protein, a plant sugar, or plant fat. In some cases, the plant protein comprises a soy protein. In some cases, the soy protein is a storage protein comprising at least one of 7s (e.g., P-conglycinin) and I ls (e.g., glycinin).

[0058] In some cases, the recombinant protein is a casein protein. In some cases, the recombinant protein is a whey protein. In some cases, the recombinant protein is an egg white protein. In some cases, the recombinant protein is a collagen protein.

[0059] In some cases, the disclosed methods further comprise adjusting pH of the mixture after adding the coagulant. In some cases, the disclosed methods further comprise adding additional coagulant after adjusting pH of the mixture. In some cases, additional coagulant is different from the coagulant. In some cases, additional coagulant is the same as the coagulant. In some cases, the disclosed methods further comprise further adjusting pH of the mixture after adding the additional coagulant. In some cases, the coagulant or the additional coagulant is at least one of a magnesium salt, a calcium salt, glucono-delta-1 actone (GDL), or any combination thereof. In some cases, the calcium salt is at least one of calcium chloride (CaCh) or calcium sulfate (CaSC ). In some cases, the magnesium salt is at least one of magnesium chloride (MgCh) or magnesium sulfate (MgSC ).

[0060] In some cases, the coagulant is added to the mixture reach a concentration above 1 mM, above 2 mM, above 3 mM, above 4 mM, above 5 mM, above 6 mM, above 7 mM, above 8 mM, above 9 mM, above 10 mM, above 11 mM, above 12 mM, above 13 mM, above 14 mM, above 15 mM, above 16 mM, above 17 mM, above 18 mM, above 19 mM, above 20 mM, above 30 mM, above 40 mM, above 50 mM, above 60 mM, above 70 mM, above 80 mM, above 90 mM, above 100 mM, above 150 mM, or above 200 mM.

[0061] In some cases, the coagulant is added to the mixture reach a concentration below 400 mM, below 390 mM, below 380 mM, below 370 mM, below 360 mM, below 350 mM, below 340 mM, below 330 mM, below 320 mM, below 310 mM, below 300 mM, below 290 mM, below 280 mM, below 270 mM, below 260 mM, below 250 mM, below 240 mM, below 230 mM, below 220 mM, below 210 mM, or below 200 mM.

[0062] In some cases, the coagulant is added to the mixture reach a concentration between 20 and 400 mM. In some cases, the coagulant is added to the mixture reach a concentration between 20 and 350 mM. In some cases, the coagulant is added to the mixture reach a concentration between 20 and 300 mM. In some cases, the coagulant is added to the mixture reach a concentration between 20 and 250 mM. In some cases, the coagulant is added to the mixture reach a concentration between 20 and 200 mM.

[0063] In some aspects, the current disclosure provides methods for extracting one or more recombinant proteins expressed in a plant, comprising obtaining a mixture from the plant comprising water, the recombinant protein(s), and 7S and 1 IS proteins in a 7S/1 IS ratio which is smaller than the natural ratio. In some aspects, the current disclosure provides methods for extracting a recombinant casein micelle expressed in a plant, comprising obtaining a mixture from the plant comprising water, the recombinant protein, and 7S and 1 IS proteins in a 7S/1 IS ratio which is smaller than the natural ratio.

[0064] In some cases, the disclosed methods further comprise at least one of cleaning the plant material to remove dirt and foreign material; dehulling or deshelling the plant material; flaking the plant material; reducing the particle size of the plant material; extracting oil from the plant material with a hexane based solvent; desolventizing the plant material without cooking and denaturing the recombinant protein; soaking the plant material in water; or any combination thereof.

[0065] In some cases, the disclosed methods are performed below 80 °C. In some cases, the disclosed methods are performed between 0 °C and 80 °C. In some cases, the disclosed methods are performed between 0 °C and 70 °C. In some cases, the disclosed methods are performed between 0 °C and 60 °C. In some cases, the disclosed methods are performed between 0 °C and 50 °C. In some cases, the disclosed methods are performed between 0 °C and 40 °C. In some cases, the disclosed methods are performed between 0 °C and 30 °C. In some cases, the disclosed methods are performed between 10 °C and 25 °C. In some cases, the disclosed methods are performed at ambient temperature.

[0066] In some aspects, the disclosed methods further comprise filtering the supernatant. In some cases, filtering the supernatant comprising using a microfiltration (MF) membrane. In some cases, the microfiltration (MF) membrane has a pore size between 0.1 and 10 pm. In some cases, the microfiltration (MF) membrane has a pore size between 0.1 and 1 pm. In some cases, microfiltration (MF) membrane has a pore size between 1 and 10 pm. In some cases, filtering the supernatant comprising at least partially removes a plant component include soluble plant proteins, plant sugars, and minerals. In some cases, the membrane filtration comprises a diafiltration step to purify a target protein. In some cases, the diafiltration solution comprises one or more salts: magnesium chloride, sodium chloride, or calcium chloride.

[0067] In some aspects, the current disclosure provides a dairy product or dairy product substitute comprising the supernatant or a substance derived from the supernatant. In some aspects, the current disclosure provides a dairy product or dairy product substitute made using the methods disclosed herein. In some cases, the dairy product or dairy product substitute is cheese. [0068] In some aspects, the current disclosure provides the dairy product or dairy product substitute comprising a recombinant casein protein and a coagulant. In some aspects, the coagulant is in a detectable amount using a standard testing method for indicating the presence of the coagulant in the dairy product or dairy product substitute. In some cases, the coagulant detectable in the dairy product or dairy product substitute is a magnesium salt, for example, at least one of magnesium chloride (MgC12) or magnesium sulfate (MgSO4). In some cases, the coagulant detectable in the dairy product or dairy product substitute is a calcium salt, for example, at least one of calcium chloride (CaC12) or calcium sulfate (CaSO4). In some cases, the coagulant detectable in the dairy product or dairy product substitute is glucono-delta-lactone (GDL).

[0069] In some cases, the coagulant detectable in the dairy product or dairy product substitute is less than 0.1% (w/w) the cheese product. In some cases, the coagulant detectable in the dairy product or dairy product substitute is less than 0.05% (w/w) the cheese product. In some cases, the coagulant detectable in the dairy product or dairy product substitute is less than 0.01% (w/w) the cheese product. In some cases, the coagulant detectable in the dairy product or dairy product substitute is less than 0.005% (w/w) the cheese product. In some cases, the coagulant detectable in the dairy product or dairy product substitute is less than 0.001% (w/w) the cheese product. In some cases, the coagulant detectable in the dairy product or dairy product substitute is more than 0.0005% (w/w) the cheese product. In some cases, the coagulant detectable in the dairy product or dairy product substitute is more than 0.0001% (w/w) the cheese product.

[0070] In some cases, the dairy product or dairy product substitute provided herein has a total fat content below 1%. In some cases, the dairy product or dairy product substitute provided herein has a total fat content below 0.1%. In some cases, the dairy product or dairy product substitute provided herein has a total fat content below 0.01%. In some cases, the dairy product or dairy product substitute provided herein has no detectable amount of animal fat. In some cases, the dairy product or dairy product substitute provided herein has no detectable amount of lactose. In some cases, the dairy product or dairy product substitute provided herein has no detectable amount of animal hormone, including, for example, at least one of estrogens, progesterone, corticoid, and androgen. In some cases, the dairy product or dairy product substitute provided herein is at least one of milk, cheese, yogurt, ghee, or butter. In some cases, the casein micelles derived from plants confer on the composition one or more characteristics of a dairy product selected from the group consisting of: taste, flavor, aroma, appearance, mouthfeel, density, structure, texture, elasticity, springiness, coagulation, binding, leavening, aeration, foaming, creaminess, and emulsification. In some cases, the composition is homogenous. In some cases, the composition is substantially homogenous.

[0071] Some aspects of the disclosure provide a composition comprising milk solids and soy proteins; wherein the soy proteins comprise glycinin and conglycinin, wherein the ratio of glycinin/conglycinin is elevated compared to a naturally occurring ratio, wherein the elevated glycinin/conglycinin ratio leads to better curd formation when the composition is subject to a curd forming condition. Some aspects of the disclosure provide a composition comprising milk solids and soy proteins; wherein the soy proteins comprise glycinin and conglycinin, wherein the ratio of glycinin/conglycinin is reduced compared to a naturally occurring ratio, wherein the reduced glycinin/conglycinin ratio leads to better curd formation when the composition is subject to a curd forming condition. In some instances, the weight ratio of the milk solids and the soy proteins is at least 60:40. In some instances, the weight ratio of the milk solids and the soy proteins is at least 65:35. In some instances, the weight ratio of the milk solids and the soy proteins is at least 70:30.

[0072] In some instances, the weight ratio of the milk solids and the soy proteins is at least 75:25.

[0073] In some instances, the weight ratio of the milk solids and the soy proteins is at least 80:20. In some instances, the weight ratio of the milk solids and the soy proteins is at most 65:35. In some instances, the weight ratio of the milk solids and the soy proteins is at most 70:30.

[0074] In some instances, the weight ratio of the milk solids and the soy proteins is at most 75:25.

[0075] In some instances, the weight ratio of the milk solids and the soy proteins is at most 80:20.

[0076] In some instances, the naturally occurring ratio is between 0.5-1.3.

[0077] In some instances UHT milk is used. In some instances raw milk is used. In some instance vat pasteurized milk is used.

BRIEF DESCRIPTION OF THE DRAWINGS

[0078] FIG. 1 A is a flow chart showing a non-limiting example of making soymilk enriched with glycinin or conglycinin.

[0079] FIG. IB shows a flow chart showing a non-limiting example of making cheese using soy proteins.

[0080] FIG. 2 shows a Sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) of enriched soy fractions.

[0081] FIG. 3 shows images of soy augmented cheese samples. [0082] FIG. 4 is a SDS-PAGE showing the results from Example 3, where conglycinin subunits a and a’ are reduced in the final retentate while glycinin subunits A 1,2, 4 and Bl, 2, 4 remain the same compared to soy dairy blend.

[0083] FIG. 5 shows the experimental result in Example 3, where curd was formed in final retentate but not in control.

[0084] FIG. 6 is a flow chart illustrating the process in Example 4A.

[0085] FIG. 7 shows SDS-PAGE of samples produced from the process described in Example 4A and shown in FIG. 6. Curd and whey samples from process derivatives analyzed. The SDS- PAGE result shows the reduction of conglycinin and glycinin subunits between the dairy and soy blend and the final retentate.

[0086] FIG.8 is a flow chart illustrating the rennet process to form a curd used in Example 4A and 4B.

[0087] FIG. 9 shows a curds formed in skim milk (control) curd (left), curd of supernatant (center), and curd of final retentate (right).

[0088] FIG. 10 shows a flow chart illustrating the process in Example 4B.

[0089] FIG. 11 shows SDS-PAGE result of samples from Example 4B using Calcium chloride to coagulate proteins and fat in the dairy and soy blend, using the process described in FIG. 8.

[0090] FIG. 12 shows from left to right: curd of supernatant (L), curd of final retentate (R) produced in Example 4B, using the process described in FIG. 8.

[0091] FIG. 13 shows MgCh concentrations at 20, 40, 60, 80, and 200 mM do not cause precipitation in skim milk.

[0092] FIG. 14 shows MgCh concentrations at 20, 40, 60, 80, and 200 mM cause precipitation in soymilk, demonstrated by pellet formation after centrifugation and solution turning clear.

[0093] FIG. 15 shows MgCh causes precipitation in a mixture of soymilk and skim milk, in a similar manner as in pure soymilk without skim milk, while the solutions do not decrease in opaqueness, suggesting only soy proteins are precipitated while milk proteins remain soluble. [0094] FIG. 16 shows MgCh concentrations at 400 mM, 800 mM, 1.2 M, 1.6 M, and 2.0 M cause precipitation in bovine skim milk.

[0095] FIG. 17 shows glycinin/conglycinin separation procedure used in Example 6, producing a glycinin fraction enriched with glycinin and a conglycinin enriched in conglycinin.

[0096] FIG. 18 shows the process of making cheese using soy fractionations in Example 6.

[0097] FIG. 19 shows the procedure used for making cheese in Example 6.

[0098] FIG. 20 shows cheese curds formation status for different fractions in Example 6.

[0099] FIG. 21 shows curd dry solids/casein ratio for different fractions in Example 6. [0100] FIG. 22 shows the total solids of the curds were measured for each curd produced in Example 6.

[0101] FIG. 23 shows a summary of data in Example 6.

[0102] FIG. 24 shows curds made from glycinin enriched soy fractions at 9g/l and 18g/l mixed with milk.

[0103] FIG. 25 shows the sequence map for plasmid pMOZ1066.

[0104] FIG. 26 shows an image from a transmission electron microscope of recombinant casein micelles decorated with anti-beta-casein-bound gold particles that were extracted from a Glycine max seed infected with pMOZ1066 expression plasmid.

[0105] FIG. 27 shows elasticity measurements for different curds in Example 6.

[0106] FIG. 28 shows curd elasticity % return to initial height for different curds in Example 6. [0107] FIG. 29 shows meltability for different curds, initial and cooked, in Example 6.

DETAILED DESCRIPTION

[0108] The following embodiments are described in sufficient detail to enable those skilled in the art to make and use the invention. It is to be understood that other embodiments would be evident based on the present disclosure, and that system, process, or mechanical changes can be made without departing from the scope of an embodiment of the present disclosure.

[0109] In the following description, numerous specific details are given to provide a thorough understanding of the invention. However, it will be apparent that the invention can be practiced without these specific details. In order to avoid obscuring an embodiment of the present disclosure, some well-known techniques, system configurations, and process steps are not disclosed in detail. Throughout this disclosure, various publications, patents and published patent specifications are referenced by an identifying citation. The disclosures of these publications, patents and published patent specifications are hereby incorporated by reference into the present disclosure.

Definitions

[0110] These and other valuable aspects of the embodiments of the present disclosure consequently further the state of the technology to at least the next level. While the disclosure has been described in conjunction with a specific best mode, it is to be understood that many alternatives, modifications, and variations will be apparent to those skilled in the art in light of the descriptions herein. Accordingly, it is intended to embrace all such alternatives, modifications, and variations that fall within the scope of the included claims. All matters set forth herein or shown in the accompanying drawings are to be interpreted in an illustrative and non-limiting sense. [OHl] As used herein, the phrases “at least one”, “one or more”, and “and/or” are open-ended expressions that are both conjunctive and disjunctive in operation. For example, each of the expressions “at least one of A, B and C”, “at least one of A, B, or C”, “one or more of A, B, and C”, “one or more of A, B, or C” and “A, B, and/or C” means A alone, B alone, C alone, A and B together, A and C together, B and C together, or A, B and C together.

[0112] As used herein, the singular forms “a”, “an” and “the” are intended to include the plural forms as well, unless the context clearly indicates otherwise. Furthermore, to the extent that the terms “including”, “includes”, “having”, “has”, “with”, or variants thereof are used in either the detailed description and/or the claims, such terms are intended to be inclusive in a manner similar to the term “comprising.”

[0113] Use of absolute or sequential terms, for example, “will,” “will not,” “shall,” “shall not,” “must,” “must not,” “first,” “initially,” “next,” “subsequently,” “before,” “after,” “lastly,” and “finally,” are not meant to limit scope of the present embodiments disclosed herein but as exemplary.

[0114] As used herein, “or” may refer to “and”, “or,” or “and/or” and may be used both exclusively and inclusively. For example, the term “A or B” may refer to “A or B”, “A but not B”, “B but not A”, and “A and B”. In some cases, context may dictate a particular meaning. [0115] Any systems, methods, software, and platforms described herein are modular and not limited to sequential steps. Accordingly, terms such as “first” and “second” do not necessarily imply priority, order of importance, or order of acts.

[0116] As used herein, the term “about” or the symbol

when referring to a number or a numerical range means that the number or numerical range referred to is an approximation within experimental variability (or within statistical experimental error), and the number or numerical range may vary from, for example, from 1% to 10% of the stated number or numerical range. Unless otherwise indicated by context, the term “about” refers to ±10% of a stated number or value.

[0117] As used herein, the term “approximately” means within an acceptable error range for the particular value as determined by one of ordinary skill in the art, which will depend in part on how the value is measured or determined, e.g., the limitations of the measurement system. For example, “approximately” can mean within 1 or more than 1 standard deviation, per the practice in the given value. Where particular values are described in the application and claims, unless otherwise stated the term “approximately” should be assumed to mean an acceptable error range for the particular value.

[0118] Where a range of values is provided, it is understood that each intervening value, to the tenth of the unit of the lower limit unless the context clearly dictates otherwise, between the upper and lower limit of that range and any other stated or intervening value in that stated range, is encompassed within the disclosure. The upper and lower limits of these smaller ranges may independently be included in the smaller ranges, and are also encompassed within the disclosure, subject to any specifically excluded limit in the stated range. Where the stated range includes one or both of the limits, ranges excluding either or both of those included limits are also included in the disclosure.

[0119] All ranges disclosed herein also encompass any and all possible sub-ranges and combinations of sub-ranges thereof. Any listed range can be recognized as sufficiently describing and enabling the same range being broken down into at least equal halves, thirds, quarters, fifths, tenths, and so forth. As a non-limiting example, each range discussed herein can be readily broken down into a lower third, middle third and upper third, and the like. All language such as “up to,” “at least,” “greater than,” “less than,” and the like include the number recited and refer to ranges which can be subsequently broken down into sub-ranges as discussed above. Finally, a range includes each individual member. Thus, for example, a group having 1-3 articles refers to groups having 1, 2, or 3 articles. Similarly, a group having 1-5 articles refers to groups having 1, 2, 3, 4, or 5 articles, and so forth.

[0120] Whenever the term “at least,” “greater than,” “greater than or equal to”, or a similar phrase precedes the first numerical value in a series of two or more numerical values, the term “at least,” “greater than,” “greater than or equal to” or similar phrase applies to each of the numerical values in that series of numerical values. For example, “at least 1, 2, or 3” is equivalent to “at least 1, at least 2, and/or at least 3.”

[0121] Whenever the term “no more than,” “less than,” “less than or equal to,” “no greater than,” “at most” or a similar phrase, precedes the first numerical value in a series of two or more numerical values, the term “no more than,” “less than,” “less than or equal to,” “no greater than,” “at most,” or similar phrase applies to each of the numerical values in that series of numerical values. For example, “less than 3, 2, or 1” is equivalent to “less than 3, less than 2, and/or less than 1.”

[0122] As used herein, the following meanings apply unless otherwise specified. The word “may” is used in a permissive sense (i.e., meaning having the potential to), rather than the mandatory sense (i.e., meaning must). The words “include”, “including”, and “includes” and the like mean including, but not limited to. The singular forms “a,” “an,” and “the” include plural referents. Thus, for example, reference to “an element” includes a combination of two or more elements, notwithstanding use of other terms and phrases for one or more elements, such as “one or more.” The phrase “at least one” includes “one”, “one or more”, “one or a plurality” and “a plurality”. The term “or” is, unless indicated otherwise, non-exclusive, i.e., encompassing both “and” and “or.” The term “any of’ between a modifier and a sequence means that the modifier modifies each member of the sequence. So, for example, the phrase “at least any of 1, 2 or 3” means “at least 1, at least 2 or at least 3”. The term "consisting essentially of refers to the inclusion of recited elements and other elements that do not materially affect the basic and novel characteristics of a claimed combination.

[0123] As used herein, a “vector” is a plasmid comprising operably linked polynucleotide sequences that facilitate expression of a coding sequence in a particular host organism (e.g., a bacterial expression vector or a plant expression vector). Polynucleotide sequences that facilitate expression in prokaryotes can include, e.g., a promoter, an enhancer, an operator, and a ribosome binding site, often along with other sequences. Eukaryotic cells can use promoters, enhancers, termination and polyadenylation signals and other sequences that are generally different from those used by prokaryotes.

[0124] As used herein, the term "casein micelles" are micelles comprising casein proteins. Examples of casein micelles are described in United States Patent Application No. 16/741,680 (Patent No. US11326176), filed on January 13, 2020, titled “Recombinant micelle and method of in vivo assembly,” and in United States Patent Application No. 17/826,021 filed on May 26, 2022, both incorporated herein by reference in its entirety. Recombinant casein micelles can be made in vivo or in vitro using the methods described therein. United States Patent Application US17/826,021 (United States Patent Application US20220290167A1), titled “Recombinant micelle and method of in vivo assembly” teaches vectors and sequences for making recombinant casein proteins and micelles, which is incorporated herein by reference in its entirety.

[0125] As used herein, the term “milk” means a liquid composition that contains soluble casein micelles and where the weight of soluble casein micelles is equal to or greater than 1% of the total protein weight in the composition.

[0126] As used herein, the term “cheese curd” is a solid or semi-solid mass made by gelating, coagulating, or curdling milk.

[0127] As used herein, the term “cheese” is a food made from cheese curds.

[0128] U.S. Patent No. 11457649 describes a substitute dairy food, and U.S. Patent Application No. 16/862,011 (Publication No. US20210010017A1) describes food compositions comprising a milk protein, both of which are incorporated herein by reference in their entirety.

[0129] As used herein, the term “7s” means P-conglycinin, or another member of the 7s globulin family of proteins from soybean or other plant species.

[0130] As used herein, the term “I ls” means glycinin, or another member of the I ls globulin family of proteins from soybean or other plant species. [0131] As used herein, the term “dairy characteristic” means a characteristic selected from one of the following characteristics of a dairy food: adhesiveness, airiness, appearance, aroma, binding, chewdown, chewiness, coagulation, cohesiveness, compactness, creaminess, crispiness, crumbliness, density, elasticity, emulsification, fattiness, firmness, flavor, foaminess, graininess, greasiness, hardness, handling, juiciness, leavening, mouthcoating, mouthfeel, richness, roughness, slipperiness on tongue, smoothness, springiness, structure, taste, tenderness, texture, thickness, uniformity, and wetness.

[0132] Where a definition or use of a term in an incorporated reference is inconsistent or contrary to the definition of that term provided herein, the definition of that term provided herein applies and the definition of that term in the reference does not apply.

[0133] As used herein, the term “recombinant” refers to nucleic acids or proteins formed by laboratory methods of genetic recombination (e.g., molecular cloning) to bring together genetic material from multiple sources, creating sequences that would otherwise not be found in the genome. Recombinant proteins may be expressed in vivo in various types of host cells, including plant cells, bacterial cells, fungal cells, avian cells, and mammalian cells. Recombinant proteins may also be generated in vitro. As used herein, the term “tagged protein” refers to a recombinant protein that includes additional peptides that are not part of the native protein and that remain after post-translational processing.

[0134] As used herein, the term “milk solids” refers to the powder that would be left after milk is dried out and the water is removed.

[0135] As used herein, the phrase “essentially free of’ is used to indicate the indicated component, if present, is present in an amount that does not contribute, or contributes only in a de minimus fashion, to the properties of the composition. In various embodiments, where a composition is essentially free of a particular component, the component is present in less than a functional amount. In various embodiments, the component may be present in trace amounts. Particular limits will vary depending on the nature of the component, but may be, for example, selected from less than 10% by weight, less than 9% by weight, less than 8% by weight, less than 7% by weight, less than 6% by weight, less than 5% by weight, less than 4% by weight, less than 3% by weight, less than 2% by weight, less than 1% by weight, less than 0.5% by weight, less than 0.1% by weight, or less than 0.05% by weight, or less than 0.01% by weight.

[0136] As used herein, the term "stably expressed" refers to expression and accumulation of a protein in a plant cell over time. As an example, a recombinant protein may accumulate because it is not degraded by endogenous plant proteases. As a further example, a recombinant protein is considered to be stably expressed in a plant if it is present in the plant in an amount of 1% or higher per total protein weight of soluble protein extractable from the plant. [0137] As used herein, the term “a detectable amount” refers to an amount of a composition (e.g., a molecule) that can be detected using the most sensitive analytical techniques up to date, including for example, liquid chromatography methods (e.g., reverse phase HPLC, size exclusion, normal phase chromatography), mass spectrometry (e.g., electrospray tandem mass spectrometry, and electrospray FT-ICR mass spectrometry), or a combination of analytical techniques (e.g., liquid chromatography-tandem mass spectrometry (LC- MS/MS)). In some cases, a detectable amount is at a concentration above 10'² mol/L, 10'³ mol/L, 10'⁴ mol/L, 10'⁵ mol/L, 1 O'⁶ mol/L, 1 O'⁷ mol/L, 1 O'⁸ mol/L, 1 O'⁹ mol/L, or 1 O'¹⁰ mol/L.

[0138] As used herein, the term “naturally occurring” means without genetic modification. For example, a naturally occurring ratio of two plant proteins means a ratio of the two plant proteins found in plant (e.g., plant seed), where the plant is not genetically modified to manipulate the expression levels of the two proteins.

[0139] As used herein, the term “recombinant” refers to nucleic acids or proteins formed by laboratory methods of genetic recombination (e.g., molecular cloning) to bring together genetic material from multiple sources, creating sequences that would otherwise not be found in the genome. Recombinant proteins may be expressed in vivo in various types of host cells, including plant cells, bacterial cells, fungal cells, avian cells, and mammalian cells. Recombinant proteins may also be generated in vitro. As used herein, the term “tagged protein” refers to a recombinant protein that includes additional peptides that are not part of the native protein and that remain after post-translational processing.

[0140] Definition of standard chemistry terms may be found in reference works, including but not limited to, Carey and Sundberg “Advanced Organic Chemistry 4th Ed.” Vols. A (2000) and B (2001), Plenum Press, New York.

[0141] As used herein, the term “homogenous” means of uniform structure or composition throughout, such that individual components (e.g., probiotics, particles) cannot be separately observed with naked eye.

[0142] As used herein, the term "plant" includes whole plant, plant organ, plant tissues, and plant cell and progeny of same, but is not limited to angiospems and gymnosperms such as Arabidopsis, potato, tomato, tobacco, alfalfa, lemice, carrot, strawberry, sugarbeet, cassava, sweet potato, soybean, lima bean, pea, chick pea, maize (com), turf grass, wheat, rice, barley, sorghum, oat, oak, eucalyptus, walnut, palm and duckweed a well as fem and moss. Thus, a plant may be a monocot, a dicot, a vascular plant reproduced from spores such as fem or a nonvascular plant such as moss, liverwort, homwort and algae. The term "plant," as used herein, also encompasses plant cells, seeds, plant progeny, propagule whether generated sexually or asexually, and descendants of any of these, such as cuttings or seed. Plant cells include suspension cultures, callus, embryos, meristematic regions, callus tissue, leaves, roots, shoots, gametophytes, sporophytes, pollen, seeds and microspores. Plants may be at various stages of maturity and may be grown in liquid or solid culture, or in soil or suitable media in pots, greenhouses or fields. As used herein, the term “plant protein” refers to a protein that is at least 70% homologous to a protein that naturally occurs in a plant.

[0143] As used herein, the term “dicof ’ refers to a flowering plant whose embryos have two seed leaves or cotyledons. Examples of dicots include Arabidopsis, tobacco, tomato, potato, sweet potato, cassava, alfalfa, lima bean, pea, chick pea, soybean, carrot, strawberry, lettuce, oak, maple, walnut, rose, mint, squash, daisy, quinoa, buckwheat, mung bean, cow pea, lentil, lupin, peanut, fava bean, French beans, mustard, or cactus.

[0144] As used herein, the term “monocof ’ refers to a flowering plant whose embryos have one cotyledon or seed leaf. Examples of monocots include turf grass, maize (corn), rice, oat, wheat, barley, sorghum, orchid, iris, lily, onion, palm, and duckweed.

[0145] As used herein, the term "transgenic plant" means a plant that has been transformed with one or more exogenous nucleic acids. "Transformation" refers to a process by which a nucleic acid is stably integrated into the genome of a plant cell. "Stably transformed" refers to the permanent, or non-transient, retention, expression, or a combination thereof of a polynucleotide in and by a cell genome. A stably integrated polynucleotide is one that is a fixture within a transformed cell genome and can be replicated and propagated through successive progeny of the cell or resultant transformed plant. Transformation can occur under natural or artificial conditions using various methods. Transformation can rely on any method for the insertion of nucleic acid sequences into a prokaryotic or eukaryotic host cell, including Agrobacterium-mediated transformation as illustrated in U.S. Pat. Nos. 5,159,135; 5,824,877; 5,591,616 and 6,384,301, all of which are incorporated herein by reference in its entirety. Methods for plant transformation also include microprojectile bombardment as illustrated in U.S. Pat. Nos. 5,015,580; 5,550,318; 5,538,880; 6,153,812; 6,160,208; 6,288,312 and 6,399,861, all of which are incorporated herein by reference in its entirety. Recipient cells for the plant transformation include meristem cells, callus, immature embryos, hypocotyls explants, cotyledon explants, leaf explants, and gametic cells such as microspores, pollen, sperm and egg cells, and any cell from which a fertile plant can be regenerated, as described in U.S. Pat. Nos. 6,194,636; 6,232,526; 6,541,682 and 6,603,061 and U.S. Patent Application publication US 2004/0216189 Al, all of which are incorporated herein by reference in its entirety.

[0146] As used herein, the term “in-vitro” means outside a living organism. [0147] As used herein, the term “fusion protein” refers to a protein comprising at least two constituent proteins that are encoded by separate genes, and that have been joined so that they are transcribed and translated as a single polypeptide.

[0148] Preparation of soybean milk.

[0149] Soybean milk can be prepared from soybeans using known methods, including these two methods:

[0150] Method 1. The beans are soaked in water at 4°C for 24 hr, dehulled and then the milk extracted by warm tap water (40°C) in the ratio of 1 :3 using a blender. It is filtered through cheese cloth to remove the insoluble residue.

[0151] Method 2. The beans are first steam heated to 60°C, then milled to produce soy flour. Soymilk is obtained by mixing soy -fl our with water in the ratio of 1 :8 blended and filtered through cheese cloth.

[0152] Rennet clotting time.

[0153] Rennet is a complex set of enzymes originally produced in the stomachs of ruminant mammals. Chymosin is the key protease enzyme that curdles casein in milk. In addition to chymosin, rennet contains other enzymes, including pepsin and a lipase. Many kinds of rennet can be used, including from animal, vegetable, and fermentation sources. Rennet clotting time can be determined visibly at 30°C in a shaker water bath, using 10 ml milk and 0.15% of calf rennet.

[0154] Curd firmness.

[0155] A milk sample (500 ml) is clotted at 30°C in a 1 -litre beaker by the addition of 0.15% calf rennet and left standing for 1 hr for complete clotting. A round aluminium plate (4 cm) with four pointed-end stands (0.2 x 10 cm) was placed on the surface of the curd. Weights were added to drive the stands into the curd and the results expressed numerically as follows: 30-35 g firm, 20- 25 soft, 10-15 very soft, less than 10 g, weak curd.

[0156] In some aspects, current disclosure provides a composition, comprising casein micelles; and a soy ingredient comprising at least one of 7s (e.g., P-conglycinin) and I ls (e.g., glycinin), wherein the ratio of 7s/l Is is lower than a naturally occurring ratio. In some instances, the lowered ratio of 7s/l Is is achieved by decreasing the amount of 7s (e.g., P-conglycinin) in the soy ingredient, for example, by using RNAi or CRISPR-Cas9 to knock out or knock down gene expression. In some instances, the lowered ratio of 7s/l Is is achieved by increasing the amount of 1 Is (e.g., glycinin) in the soy ingredient, for example, by overexpressing I ls.

[0157] In some cases, the naturally occurring ratio of 7s/l Is is 0.5. In some cases, the naturally occurring ratio of 7s/l Is is 0.6. In some cases, the naturally occurring ratio of 7s/l Is is 0.7. In some cases, the naturally occurring ratio of 7s/l Is is 0.75. In some cases, the naturally occurring ratio of 7s/l Is is 0.8. In some cases, the naturally occurring ratio of 7s/l Is is 0.9. In some cases, the naturally occurring ratio of 7s/l Is is 1. In some cases, the naturally occurring ratio of 7s/l Is is 1.1. In some cases, the naturally occurring ratio of 7s/l Is is 1.2 In some cases, the naturally occurring ratio of 7s/l Is is 1.3.

[0158] In some cases, the ratio of 7s/l Is is less than 0.4. In some cases, the ratio of 7s/l Is is less than 0.3. In some cases, the ratio of 7s/l Is is less than 0.25. In some cases, the ratio of 7s/l Is is less than 0.2. In some cases, the ratio of 7s/l Is is less than 0.1. In some cases, the ratio of 7s/l Is is 0 or close to 0, wherein the soy ingredient does not comprise a detectable amount of 7s (e.g., P-conglycinin).

[0159] In some cases, the soy ingredient is in the form of soy protein isolate. In some cases, the soy ingredient is in the form of soymilk.

[0160] In some cases, the source of the casein micelles is from an animal, for example a mammal, for example, a human. In some cases, the mammal is a ruminant. In some cases, the ruminant is bovine, sheep, or goat. In some cases, source of the casein micelles is from a genetically modified microorganism, for example, yeast, fungi, or bacteria. In some cases, the source of the casein micelles is from a genetically modified plant. In some cases, the genetically modified plant is a soybean plant.

[0161] In some cases, casein protein is less than 50% (w/w) of total protein in the composition. In some cases, casein protein is less than 60% (w/w) of total protein in the composition. In some cases, casein protein is less than 70% (w/w) of total protein in the composition. In some cases, casein protein is less than 80% (w/w) of total protein in the composition. In some cases, casein protein is less than 90%(w/w) of total protein in the composition.

[0162] In some cases, soy protein is more than 30% (w/w) of total protein in the composition. In some cases, soy protein is more than 40% (w/w) of total protein in the composition. In some cases, soy protein is more than 50% (w/w) of total protein in the composition. In some cases, soy protein is more than 60% (w/w) of total protein in the composition. In some cases, soy protein is more than 70% (w/w) of total protein in the composition. In some cases, soy protein is more than 80% (w/w) of total protein in the composition. In some cases, soy protein is more than 90% (w/w) of total protein in the composition.

[0163] In some cases, the composition forms a cheese curd in a cheese making process. In some cases, the composition forms a firm cheese curd in a cheese making process. In some cases, the composition forms a firm cheese curd in a cheese making process within 15 minutes in the cheese making process. In some cases, the composition forms a firm cheese curd in a cheese making process within 30 minutes in the cheese making process. In some cases, the composition forms a firm cheese curd in a cheese making process within 60 minutes in the cheese making process. In some cases, the composition forms a firm cheese curd in a cheese making process within 100 minutes in the cheese making process.

[0164] In some cases, the composition forms a soft cheese curd in a cheese making process. In some cases, the composition forms a soft cheese curd within 15 minutes in the cheese making process. In some cases, the composition forms a soft cheese curd within 30 minutes in the cheese making process. In some cases, the composition forms a soft cheese curd within 60 minutes in the cheese making process. In some cases, the composition forms a soft cheese curd within 100 minutes in the cheese making process.

[0165] In some cases, the cheese making process is performed at pH between 6.0-7.0. In some cases, the cheese making process is performed at pH between 6.0-6.5. In some cases, the cheese making process is performed at pH between 6.5-7.0. In some cases, the cheese making process comprises adding rennet to the composition to cause the casein micelle to precipitate.

[0166] In some aspects, the current disclosure provides a cheese product comprising the composition disclosed herein. In some aspects, the current disclosure provides a method of making cheese, comprising providing the composition disclosed herein, and adding rennet to the composition to cause the casein micelles to precipitate.

[0167] In some aspects, the current disclosure provides compositions, methods and systems for making cheese using a modified soy ingredient. In some aspects, the modified soy ingredient comprises a non-detectable or reduced level of an anti -nutritional or toxin in soybean (for example, including trypsin inhibitors, phytic acid, lectins, or soybean toxin), compared to a natural occurring level found in non-modified soybeans. For example, the non-detectable or reduced levels of anti -nutritional or toxin can be achieved by knocking out or down one or more of the following genes: anti-nutritional/toxins in soybean, including trypsin inhibitors, phytic acid, lectins, soybean toxin.

[0168] In some aspects, the current disclosure provides compositions, methods and systems for making cheese using one or more seed storage proteins (e.g., legumin, vicilins, prolamin gliadin, etc.).

[0169] In some aspects, the current disclosure also provides food compositions comprising the composition disclosed herein or made using the methods disclosed herein. Contemplated food compositions include dairy products or products that resembles a dairy product (i.e., dairy product substitutes). The term "dairy product" as used herein refers to milk (e.g., whole milk (at least 3.25% milk fat), partly skimmed milk (from 1% to 2% milk fat), skim milk (less than 0.2% milk fat), cooking milk, condensed milk, flavored milk, goat milk, sheep milk, dried milk, evaporated milk, milk foam), and products derived from milk, including but not limited to yogurt (e.g., whole milk yogurt (at least 6 grams of fat per 170 g), low-fat yogurt (between 2 and 5 grams of fat per 170 g), nonfat yogurt (0.5 grams or less of fat per 170 g), greek yogurt (strained yogurt with whey removed), whipped yogurt, goat milk yogurt, Labneh (labne), sheep milk yogurt, yogurt drinks (e.g., whole milk Kefir, low-fat milk Kefir), Lassi), cheese (e.g., whey cheese such as ricotta; pasta filata cheese such as mozzarella; semi-soft cheese such as Havarti and Muenster; medium-hard cheese such as Swiss and Jarlsberg; hard cheese such as Cheddar and Parmesan; washed curd cheese such as Colby and Monterey Jack; soft ripened cheese such as Brie and Camembert; fresh cheese such as cottage cheese, feta cheese, cream cheese, and curd; processed cheese; processed cheese food; processed cheese product; processed cheese spread; enzyme-modulated cheese; cold-pack cheese), dairy-based sauces (e.g., fresh, frozen, refrigerated, or shelf stable), dairy spreads (e.g., low-fat spread, low-fat butter), cream (e.g., dry cream, heavy cream, light cream, whipping cream, half-and-half, coffee whitener, coffee creamer, sour cream, creme fraiche), frozen confections (e.g., ice cream, smoothie, milk shake, frozen yogurt, sundae, gelato, custard), dairy desserts (e.g., fresh, refrigerated, or frozen), ghee, butter (e.g., whipped butter, cultured butter), dairy powders (e.g., whole milk powder, skim milk powder, fat-filled milk powder (i.e., milk powder comprising plant fat in place of all or some animal fat), infant formula, milk protein concentrate (i.e., protein content of at least 80% by weight), milk protein isolate (i.e., protein content of at least 90% by weight), whey protein concentrate, whey protein isolate, demineralized whey protein concentrate, demineralized whey protein concentrate, .beta.-lactoglobulin concentrate, .beta.-lactoglobulin isolate, .alpha.- lactalbumin concentrate, .alpha.-lactalbumin isolate, glycomacropeptide concentrate, glycomacropeptide isolate, casein concentrate, casein isolate, nutritional supplements, texturizing blends, flavoring blends, coloring blends), ready-to-drink or ready-to-mix products (e.g., fresh, refrigerated, or shelf stable dairy protein beverages, weight loss beverages, nutritional beverages, sports recovery beverages, and energy drinks), puddings, gels, chewables, crisps, and bars. The term "food product substitute" (e.g., "dairy product substitute") as used herein refers to a food product that resembles a conventional food product (e.g., can be used in place of the conventional food product). Such resemblance can be due to any physical, chemical, or functional attribute. In some embodiments, the resemblance of the food product provided herein to a conventional food product is due to a physical attribute. Non-limiting examples of physical attributes include color, shape, mechanical characteristics (e.g., hardness, G' storage modulus value, shape retention, cohesion, texture (i.e., mechanical characteristics that are correlated with sensory perceptions (e.g., mouthfeel, fattiness, creaminess, homogenization, richness, smoothness, thickness), viscosity, and crystallinity. In some embodiments, the resemblance of the food product provided herein and a conventional food product is due to a chemical/biological attribute. Non-limiting examples of chemical attributes include nutrient content (e.g., type and/or amount of amino acids (e.g., PDCAAS score), type and/or amount of lipids, type and/or amount of carbohydrates, type and/or amount of minerals, type and/or amount of vitamins), pH, digestibility, shelf-life, hunger and/or satiety regulation, taste, and aroma. In some embodiments, the resemblance of the food product provided herein to a conventional food product is due to a functional attribute. Nonlimiting examples of functional attributes include gelling/agglutination behavior (e.g., gelling capacity (i.e., time required to form a gel (i.e., a protein network with spaces filled with solvent linked by hydrogen bonds to the protein molecules) of maximal strength in response to a physical and/or chemical condition (e.g., agitation, temperature, pH, ionic strength, protein concentration, sugar concentration, ionic strength)), agglutination capacity (i.e., capacity to form a precipitate (i.e., a tight protein network based on strong interactions between protein molecules and exclusion of solvent) in response to a physical and/or chemical condition), gel strength (i.e., strength of gel formed, measured in force/unit area (e.g., pascal (Pa))), water holding capacity upon gelling, syneresis upon gelling (i.e., water weeping over time)), foaming behavior (e.g., foaming capacity (i.e., amount of air held in response to a physical and/or chemical condition), foam stability (i.e., half-life of foam formed in response to a physical and/or chemical condition), foam seep), thickening capacity, use versatility (i.e., ability to use the food product in a variety of manners and/or to derive a diversity of other compositions from the food product; e.g., ability to produce food products that resemble milk derivative products such as yoghurt, cheese, cream, ghee, and butter), and ability to form protein dimers.

[0170] In some aspects, the current disclosure provides methods for extracting a recombinant protein expressed in a plant, comprising obtaining a mixture comprising water, the recombinant protein, and a component from the plant; adding a coagulant to the mixture to cause at least a portion of the plant component to coagulant; and extracting supernatant from the mixture, wherein the supernatant comprises the recombinant protein. In some cases, the methods are performed in sequence. In some cases, the mixture comprises a slurry made by blending the plant in water. In some cases, the disclosed methods further comprise separating insoluble plant components from the slurry to produce a plant-based milk before adding the coagulant to the mixture.

[0171] In some cases, the plant is a legume. In some cases, the plant is at least one of alfalfa, cassava, cotton, cowpea, maize, pea, peanut, rice, sesame, sorghum, soybean, or yam. In some cases, the plant component comprises a plant protein, a plant sugar, or plant fat. In some cases, the plant protein comprises a soy protein. In some cases, the soy protein is a storage protein comprising at least one of 7s (e.g., P-conglycinin) and I ls (e.g., glycinin).

[0172] In some cases, the recombinant protein is a casein protein. In some cases, the recombinant protein is one or more casein proteins. In some cases, the recombinant protein is a whey protein. In some cases, the recombinant protein is an egg white protein. In some cases, the recombinant protein is a collagen protein. In some cases the recombinant protein is a casein micelle. In some cases the recombinant protein is a fusion protein. In some cases the recombinant protein is one or more fusion proteins.

[0173] In some cases, the disclosed methods further comprise adjusting pH of the mixture after adding the coagulant. In some cases, the disclosed methods further comprise adding additional coagulant after adjusting pH of the mixture. In some cases, additional coagulant is different from the coagulant. In some cases, additional coagulant is the same as the coagulant. In some cases, the disclosed methods further comprise further adjusting pH of the mixture after adding the additional coagulant.

[0174] In some cases, the coagulant or the additional coagulant is at least one of a magnesium salt, a calcium salt, glucono-delta-lactone (GDL), or any combination thereof. In some cases, the calcium salt is at least one of calcium chloride (CaCh) or calcium sulfate (CaSC ). In some cases, the magnesium salt is at least one of magnesium chloride (MgCh) or magnesium sulfate (MgSC ). In some cases, the coagulant is added to the mixture reach a concentration between 5 and 200 mM. In some cases, the coagulant is added to the mixture reach a concentration between 10 and 100 mM. In some cases, the coagulant is added to the mixture reach a concentration between 20 and 80 mM. In some cases, the coagulant is added to the mixture reach a concentration between 30 and 70 mM. In some cases, the coagulant is added to the mixture reach a concentration between 30 and 50 mM.

[0175] In some cases, the disclosed methods further comprise at least one of cleaning the plant material to remove dirt and foreign material; dehulling or deshelling the plant material; flaking the plant material; reducing the particle size of the plant material; extracting oil from the plant material with a hexane based solvent; desolventizing the plant material without cooking and denaturing the recombinant protein; soaking the plant material in water; or any combination thereof.

[0176] In some cases, the disclosed methods are performed below 80 °C. In some cases, the disclosed methods are performed between 0 °C and 80 °C. In some cases, the disclosed methods are performed between 0 °C and 70 °C. In some cases, the disclosed methods are performed between 0 °C and 60 °C. In some cases, the disclosed methods are performed between 0 °C and 50 °C. In some cases, the disclosed methods are performed between 0 °C and 40 °C. In some cases, the disclosed methods are performed between 0 °C and 30 °C. In some cases, the disclosed methods are performed between 10 °C and 25 °C. In some cases, the disclosed methods are performed at ambient temperature. [0177] In some cases, this disclosure provides methods for improving mammalian glycan-related functionality of a protein, particularly compared to an unmodified version of the protein. The improvement can be, in some cases, at least 1.25-fold, 1.5-fold, 1.75-fold, 2-fold, 3-fold, 4-fold, 5-fold, 6-fold, 7-fold, 8-fold or more. In some cases, the glycan-related functionality is increased solubility (e.g., increased solubility of the modified protein compared to unmodified protein) of the protein (e.g., casein). In some cases, the glycan-related functionality is increased stability. [0178] In some aspects, the disclosed methods further comprise filtering the supernatant. In some cases, filtering the supernatant comprising using a microfiltration (MF) membrane. In some cases, the microfiltration (MF) membrane has a pore size between 0.1 and 10 pm. In some cases, the microfiltration (MF) membrane has a pore size between 0.1 and 1 pm. In some cases, microfiltration (MF) membrane has a pore size between 1 and 10 pm. In some cases, filtering the supernatant comprising at least partially removes a plant component include soluble plant proteins, plant sugars, and minerals. In some case, the membrane filtration comprises a diafiltration step to purify a target protein. In some cases, the diafiltration solution comprises one or more salts: magnesium chloride, sodium chloride, or calcium chloride.

[0179] In some aspects, the current disclosure provides a dairy product or dairy product substitute comprising the supernatant or a substance derived from the supernatant. In some aspects, the current disclosure provides a dairy product or dairy product substitute made using the methods disclosed herein. In some cases, the dairy product or dairy product substitute is cheese. [0180] In some aspects, the current disclosure provides the dairy product or dairy product substitute comprising a recombinant casein protein and a coagulant. In some aspects, the coagulant is in a detectable amount using a standard testing method for indicating the presence of the coagulant in the dairy product or dairy product substitute. In some cases, the coagulant detectable in the dairy product or dairy product substitute is a magnesium salt, for example, at least one of magnesium chloride (MgCh) or magnesium sulfate (MgSC ). In some cases, the coagulant detectable in the dairy product or dairy product substitute is a calcium salt, for example, at least one of calcium chloride (CaCh) or calcium sulfate (CaSC ). In some cases, the coagulant detectable in the dairy product or dairy product substitute is glucono-delta-lactone (GDL).

[0181] In some cases, the coagulant detectable in the dairy product or dairy product substitute is less than 0.1% (w/w) the cheese product. In some cases, the coagulant detectable in the dairy product or dairy product substitute is less than 0.05% (w/w) the cheese product. In some cases, the coagulant detectable in the dairy product or dairy product substitute is less than 0.01% (w/w) the cheese product. In some cases, the coagulant detectable in the dairy product or dairy product substitute is less than 0.005% (w/w) the cheese product. In some cases, the coagulant detectable in the dairy product or dairy product substitute is less than 0.001% (w/w) the cheese product. In some cases, the coagulant detectable in the dairy product or dairy product substitute is more than 0.0005% (w/w) the cheese product. In some cases, the coagulant detectable in the dairy product or dairy product substitute is more than 0.0001% (w/w) the cheese product.

[0182] In some cases, the dairy product or dairy product substitute provided herein has a total fat content below 1%. In some cases, the dairy product or dairy product substitute provided herein has a total fat content below 0.1%. In some cases, the dairy product or dairy product substitute provided herein has a total fat content below 0.01%. In some cases, the dairy product or dairy product substitute provided herein has no detectable amount of animal fat. In some cases, the dairy product or dairy product substitute provided herein has no detectable amount of lactose. In some cases, the dairy product or dairy product substitute provided herein has no detectable amount of animal hormone, including, for example, at least one of estrogens, progesterone, corticoid, and androgen. In some cases, the dairy product or dairy product substitute provided herein is at least one of milk, cheese, yogurt, or butter.

[0183] As used herein, ambient temperature is a colloquial expression for the typical or preferred indoor (climate-controlled) temperature to which people are generally accustomed. It represents the small range of temperatures at which the air feels neither hot nor cold, approximately 21° C. In some embodiments, ambient temperature is 25±5° C. In some embodiments, ambient temperature is 18° C. In some embodiments, ambient temperature is 19° C. In some embodiments, ambient temperature is 20° C. In some embodiments, ambient temperature is 21° C. In some embodiments, ambient temperature is 22° C. In some embodiments, ambient temperature is 23° C. In some embodiments, ambient temperature is 24° C. In some embodiments, ambient temperature is 25° C. In some embodiments, ambient temperature is 26° C. In some embodiments, ambient temperature is 27° C. In some embodiments, ambient temperature is 28° C. In some embodiments, ambient temperature is 29° C. In some embodiments, ambient temperature is 30° C.

Example 1

[0184] This non-limiting example shows adding soy proteins to milk or other compositions of casein micelles reduces the quality of casein micelle coagulation during the cheesemaking process. Increasing the ratio of glycinin to conglycinin in the soy protein enables a higher soy protein inclusion rate in curd-forming mixtures. The process is also illustrated in FIG. 1 (FIG. 1A and FIG. IB)

[0185] 1. Soymilk was produced from defatted soy white flakes with a high protein dispersibility index (PDI). The white flakes were mixed with warm, deionized (DI) water to produce a 10 wt% slurry. The pH was adjusted to pH of 8 with 2 N Sodium Hydroxide (NaOH). The slurry was mixed for 10 minutes to extract proteins, sugars, salts, and other materials from the white flake. The slurry was then centrifuged to separate the soluble soymilk (centrate) from the insoluble centrifuge cake (okara). This soymilk was the “standard” soymilk. The okara was discarded.

[0186] 2 Sodium sulfate was added to the soymilk until the sodium sulfate concentration was 30 mM. The pH was adjusted down to 6.0 with 2 N Hydrochloric Acid (HC1). The combination of lower pH and salt addition precipitated a glycinin-rich protein fraction from the soymilk. The mixture was then refrigerated overnight.

[0187] 3. The glycinin-rich precipitate was removed by centrifugation and set aside. The remaining centrate contained a majority of the conglycinin proteins and was labeled “CG” soymilk.

[0188] 4. The glycinin-rich protein precipitate was resuspended in DI water and the pH was adjusted up to 7.5 using 2 N NaOH to produce “G” soymilk.

[0189] 5. Homogenized, pasteurized skim milk was blended with one of three different soymilk types: “standard,” “CG,” or “G”. The soymilks were added at either a low or high dosage, where the low dosage was 9 grams of soy solids per liter of mixture and the high dosage was 18 grams per liter. In addition, a control sample was prepared with only skim milk and no soy protein added.

[0190] 6. The blends of skim milk and soy proteins were then subjected to a typical rennetbased cheesemaking process (ie as described in example 8).

[0191] 7 Of the seven blends tested, the only blends observed to produce a cheese curd were the control (no added soy) and both the low and high dosage “G” (glycinin-enriched) soymilk. Curds from the glycinin-enriched blends are depicted in FIG. 24. The “standard” and “CG” soymilks did not form curds.

Example 2

[0192] In this non-limiting example, the major soy proteins, glycinin and P-conglycinin, were separated into two different fractions. One fraction was enriched in glycinin, and the second fraction enriched in P-conglycinin. Milk was separately combined with the individual fractions as well as standard soymilk. The mixtures of soy and dairy milk were subjected to a rennet-based cheese process. At higher inclusions of soy, the milk augmented with the fraction enriched in glycinin produced a cheese curd, while the milks augmented with either conglycinin or standard soymilk did not.

[0193] In this experiment, soymilk was produced from defatted soy white flakes with a high protein dispersibility index (PDI). The white flakes were mixed with warm, deionized (DI) water to produce a 10 wt% slurry. The pH was adjusted to pH of 8 with 2 N Sodium Hydroxide (NaOH). The slurry was mixed for 10 minutes to extract proteins, sugars, salts, and other materials from the white flake. The slurry was then centrifuged to separate the soluble soymilk (centrate) from the insoluble centrifuge cake (okara). This soymilk is the “standard” soymilk. The okara was discarded.

[0194] Sodium sulfate was added to the soymilk until the sodium sulfate concentration was 30 mM. The pH was adjusted down to 6.0 with 2 N HC1. The combination of lower pH and salt addition precipitates the glycinin-rich protein fraction from the soymilk. The mixture was refrigerated overnight.

[0195] The glycinin-rich precipitate was removed by centrifugation and set aside. The remaining centrate contained a majority of the conglycinin proteins and was labeled as “CG” soymilk.

[0196] The glycinin-rich protein precipitate was resuspended in DI water and the pH was adjusted to 7.5 using 2 N NaOH to produce “G” soymilk.

[0197] The “G” soymilk was concentrated to a 2x concentration factor with a lOOkDa PVDF membrane in a benchtop tangential flow filtration system. The lOOkDa PVDF membrane was labeled “G membrane”. The permeate was discarded.

[0198] The concentrated “G” soymilk (retentate) was diluted with (1) volume of diafiltration DI water and concentrated to a 2x concentration factor with the G membrane. The permeate was again discarded.

[0199] The concentrated “G” soymilk (retentate) was diluted with (1) volume of diafiltration DI water and again concentrated to a 2x concentration factor with the G membrane. Diafiltration removed the dissolved proteins, sugars, minerals, and salts. The “G” soymilk was relabeled as “Washed G” and set aside. The permeate was again discarded.

[0200] With a clean, new 100 kDa PVDF membrane, the “CG” soymilk was concentrated to a 2x concentration factor using a benchtop tangential flow filtration system. This membrane was labeled “CG Membrane”.

[0201] The concentrated “CG” soymilk was washed with (1) volume of 20 mM sodium sulfate and reconcentrated to a 2x concentration factor with the CG membrane.

[0202] The concentrated “CG” soymilk was diluted with (1) volume of diafiltration DI water and concentrated to a 2x concentration factor with the CG membrane. The “CG” soymilk was relabeled as “Washed CG”.

[0203] The “Washed CG” and “standard” soymilk were diluted with DI water to the same total solids concentration as the “Washed G” soymilk.

[0204] The individual protein fractions were analyzed with sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS PAGE). The SDS-page shows the conglycinin fraction depleted in glycinin, and the glycinin fraction depleted of conglycinin. Protein levels were determined with a Millipore Bicinchoninic Acid (BCA) assay and each lane was dosed with 5 pg of protein. An image of the SDS-page described is presented in FIG. 2.

[0205] Homogenized, pasteurized skim milk was blended with one of three different soymilk types: “Standard,” “Washed CG,” or “Washed G”. The soymilks were added at a low and high dosage, where the low dosage was 4.5 grams of soy solids per liter of mixture and the high dosage was 13.5 grams per liter. A control sample was prepared with only skim milk and no soy protein added.

[0206] The blends of skim milk and soy proteins were then subjected to a rennet-based cheesemaking process. The mixture was coagulated for 50 minutes in a 37 C water bath. The control and all samples with the low dose (4.5 g/L soy solids) produced a cheese curd. At the high dose, only the “Washed G” fraction produced a cheese curd. The high dose of “Standard” and “Washed CG” did not produce a cheese curd. An image of the cheese curds is presented in FIG. 3 Example 3

[0207] This is a non-limiting example demonstrating reducing conglycinin concentrations in soy/dairy blends improves curd quality when compared to soy/dairy blends with unaltered conglycinin: glycinin ratios. In this experiment, whole soymilk and bovine skim milk were combined and treated with a solution of calcium chloride to salt out soy proteins. The insoluble soy proteins were removed via centrifugation. The supernatant was concentrated and washed with a microfiltration (MF) membrane in a benchtop tangential flow filtration system to wash out and remove additional soy proteins while retaining casein micelles. The final retentate of the filtered soy and dairy blend was subjected to a rennet-based cheese making process. The soy/dairy blend control did not produce a curd while the filtered and washed retentate did make a curd.

[0208] In this experiment, soymilk was produced from defatted soy white flakes with a high protein dispersibility index (PDI). The white flakes were mixed with warm, deionized (DI) water to produce a 10 wt% slurry. The pH was not adjusted. The slurry was mixed for 10 minutes to extract proteins, sugars, salts, and other materials from the white flake. The slurry was centrifuged at 3200 g-force for 10 minutes to separate the soluble soymilk (centrate) from the insoluble centrifuge cake (okara). This soymilk is the “standard” soymilk. A sample of the “standard” soymilk was set aside for further analysis. The okara was discarded.

[0209] Ultra-High Temperature (UHT) pasteurized and homogenized bovine skim milk was combined with the “standard” soymilk at a volumetric ratio of 1 :2 soymilk: skim milk. The pH of the soy/dairy blend was measured and recorded. This mixture was labeled as “soy/dairy blend”. [0210] A solution of 200 g/L calcium chloride was made with warm DI water and solid calcium chloride.

[0211] The 200 g/L solution of calcium chloride was added to the soy dairy blend to achieve a 10 mM calcium chloride concentration. The pH was adjusted back to the soy/dairy blend’s initial pH by adding 2 N sodium hydroxide. The calcium chloride addition and pH step was repeated twice. The final calcium chloride concentration of the solution was 30 mM. This material was called “30 mM soy/dairy blend”.

[0212] The 30 mM soy/dairy blend was heated to 50 C on a hot plate while mixing with a stir bar. The solution was stirred for 10 minutes at 50 C.

[0213] The 30 mM soy/dairy blend was transferred to centrifuge bottles and centrifuged at 3200 g-force for 10 minutes. The supernatant was separated from the pellet of insoluble proteins. The supernatant was labeled “30 mM soy/dairy supernatant”.

[0214] The 30 mM soy dairy supernatant was concentrated to a 2x concentration factor with a 0.65 micron PES membrane in a benchtop tangential flow filtration system. The permeate was discarded.

[0215] The concentrated dairy soy blend (retentate) was diluted with (1) volume of diafiltration DI water and concentrated to a 2x concentration factor with the same 0.65 micron membrane. The permeate was again discarded.

[0216] The concentrated dairy soy blend (retentate) was diluted with (1) volume of diafiltration DI water and again concentrated to a 2x concentration factor with the 0.65 micron membrane. Diafiltration removed the dissolved proteins, sugars, minerals, and salts. The permeate was again discarded. The retentate was set aside and labeled as “Final Retentate”.

[0217] The individual protein fractions were analyzed with sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS PAGE). The SDS-PAGE of the final retentate shows conglycinin is reduced from the initial soy/dairy blend. The visible glycinin subunits Al, 2, 4 and B 1,2,4 remain in the final retentate. Protein levels were determined with a Millipore Bicinchoninic Acid (BCA) assay and each lane was dosed with 5 pg of protein. An image of the SDS-page described is presented in FIG. 4.

[0218] The final retentate and the initial soy/dairy blend (control) samples were subjected to a rennet-based cheesemaking process. The mixture was coagulated for 50 minutes in a 37 C water bath. The control sample did not produce a rennet curd. The final retentate produced a rennet curd. An image of the cheese curds is presented in FIG. 5.

Example 4 A

[0219] This non-limiting example shows soybean and casein micelles were separated as an example of the separation and purification process described herein. The example demonstrates native plant proteins can be selectively removed and casein micelles purified by first removing the soybean components through a coagulation process and purification in subsequent membrane filtration. The process carried out in Example 4A is illustrated in FIG 6.

[0220] 1. Full-fat soymilk was produced from whole soybeans (glycine max). The soybeans were first ground in a food processor to flour until a fine uniform distribution was produced. The soy flour was mixed with 65 °C deionized (DI) water to produce a 10 wt% slurry. The slurry was mixed for 10 minutes to extract proteins, sugars, salts, and other materials from the soybean. The slurry was then centrifuged to separate the soluble soymilk (centrate) from the insoluble centrifuge cake (okara). This soymilk is the “Full-Fat Soymilk”. The okara was discarded.

[0221] 2. The full-fat soymilk was combined with bovine skim milk (homogenized and Ultra- high temperature (UHT) pasteurized) to produce a solution that contained equal parts casein protein and equal parts soy solids. The casein content of skim milk was assumed to be 27 g/L. A portion labeled “Dairy and soy Blend” was set aside for additional tests (SDS-PAGE, rennet process, moisture analysis, pH).

[0222] 3. A 200 g/L concentrated solution of magnesium chloride was prepared by dissolving anhydrous magnesium chloride (MgCh, CAS No. 7786-30-3) in room-temperature DI water. [0223] 4. The dairy and soy blend was heated to 20C on a heated stir plate. After the solution reached 20 °C, magnesium chloride was added to 10 mM magnesium chloride. The pH was adjusted back to 6.7 with 2 N NaOH. This process was repeated in 10 mM increments until the concentration reached 40 mM. The solution was held at 20 °C for 20 minutes after all the magnesium chloride was added to allow the proteins to coagulate.

[0224] 5. The magnesium chloride treated dairy and soy blend was centrifuged for 10 minutes at 3700 rpm. The supernatant was decanted from the solids. The solids were weighed and the moisture content measured. An aliquot of supernatant was set aside for a rennet cheese process, SDS-PAGE, moisture measurement, and pH.

[0225] 6. The supernatant was concentrated to a 2x concentration factor with a 0.65 micron polyethersulfone (PES) membrane in a benchtop tangential flow filtration system. The membrane was labeled 0.65 micron PES. The permeate was discarded.

[0226] 7. The concentrated supernatant was washed with (1) volume of DI H2O and reconcentrated to a 2x concentration factor with the 0.65 micron PES membrane.

[0227] 8. The concentrated supernatant was again washed with (1) volume of DI H2O and reconcentrated to a 2x concentration factor with the 0.65 micron PES membrane.

[0228] 9. The concentrated solution was diluted 2x with DI H2O, labeled “final retentate” and set aside for rennet process, SDS-PAGE, moisture measurement, and pH. [0229] 10. The individual protein fractions were analyzed with sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE). The SDS-PAGE shows the final retentate has reduced levels of native soy proteins when compared with the initial dairy and soy blend. Protein levels were determined with a Millipore Bicinchoninic Acid (BCA) assay and each lane was dosed with 4 pg of protein. An image of the SDS-PAGE described is presented in FIG. 7. Final retentate has a reduced soy protein concentration compared to the supernatant, suggesting membrane filtration further removed soy proteins.

[0230] 11. The final retentate, initial dairy and soy blend, supernatant, and skim milk were subjected to a rennet-based cheesemaking process. The mixtures were coagulated for 50 minutes at 32 °C. The rennet process is described in FIG. 8. The initial dairy and soy blend did not produce a curd. The supernatant, final retentate, and skim milk produced a curd. An image of the curds is presented in FIG. 9. The final retentate curd was observed to have one or more improved dairy characteristics including, having a lower moisture content, better color (more white), and had better elasticity than the supernatant curd.

Example 4B

[0231] In this non-limiting example (illustrated in FIG. 10), which is an expansion of Example 4A, soy and casein proteins are separated using calcium chloride as the coagulant instead of magnesium chloride. Other coagulating agents can be used in the separation of recombinant proteins from plant materials.

[0232] The same procedure executed in Example 1 A was repeated except calcium chloride replaced magnesium chloride at the same 40 mM concentration. The results of Example IB were similar to Example 1 A. The SDS-PAGE (shown in FIG. 11) showed reduced conglycinin and glycinin in the supernatant and final retentate than the initial dairy and soy blend. The casein proteins became functional and formed a curd in a rennet process after the coagulation and membrane filtration processes. The curd produced in the rennet process possessed similar characteristics to that of cheese made from calcium fortified skim milk. The micelles were functional and did form a curd with higher moisture, reduced meltability, and limited elasticity compared to a skim milk control curd, as shown in FIG. 12.

Example 5

[0233] In this non-limiting example, bovine skim milk, whole soymilk, and a dairy and soy blend, were each treated with different concentrations of magnesium chloride and then centrifuged.

[0234] 1. In this experiment, full-fat soymilk was produced from whole soybeans (glycine max). The soybeans were first ground in a food processor until a fine flour was produced. The soy flour was mixed with 65 °C deionized (DI) water to produce a 10 wt% slurry. The slurry was mixed for 10 minutes to extract proteins, sugars, salts, and other materials from the soybeans. The slurry was then centrifuged to separate the soluble soymilk (centrate) from the insoluble centrifuge cake (okara). This soymilk is the whole soymilk. The okara was discarded. [0235] 2. The full-fat soymilk was combined with bovine skim milk (homogenized and Ultra-high temperature (UHT) pasteurized) to produce a solution that contained equal parts casein protein and equal parts soy solids (47 vol% soymilk, 53 vol% skim milk). The casein content of skim milk was assumed to be 27 g/L.

[0236] 3. A concentrated salt solution was prepared with the anhydrous forms of magnesium chloride. The liquid coagulating agent was prepared to a concentration of 200 g/L by dissolving the salt in room -temperature DI water.

[0237] 4. The soymilk, skim milk, and dairy and soy blend samples were each split into two beakers (six total). One of the two beakers was kept at room temperature (20 °C), while the other beaker was heated to 50 °C. A 50 mL control sample was taken when the solutions reached 50 °C. The salt solutions were added in 10 mM increments and pH adjusted to 6.7 with 2 N NaOH. This process was repeated until the salt concentration reached 200 mM. A 50 mL sample was taken at 20, 40, 60, 80, and 200 mM and labeled according to the salt concentration and sample type.

[0238] 5. The 50 mL samples were centrifuged for 10 minutes at 3700 RPM. The gradient of samples were photographed. The supernatant was decanted from the solids. The solids were removed from the conical tubes with DI water and dried in an oven. The weight of the dry solids was measured and recorded. The ratio of (gram precipitated solids)/( starting solution volume) was compared between samples.

[0239] a. Skim milk: MgCh concentrations 20, 40, 60, 80 and 200 mM did not cause precipitation noticeably different than the control (skim milk without magnesium chloride). The solutions at all concentrations of magnesium chloride remain opaque, and there is no pellet formation after centrifugation (as shown in FIG. 13).

[0240] b. Soymilk: MgCh treatment changed the color of soymilk samples from opaque to a yellow-tinted transparent fluid, and produced solid pellets after centrifugation. Magnesium chloride at concentrations 60, 80 and 200 mM did not further increase the amount of precipitated solids or decrease the opaqueness of the solutions, compared to 40 mM MgCh, as shown in FIG.

14

[0241] c. Skim milk and soymilk mixture: MgCh treatment caused precipitation in a mixture of soymilk and skim milk, in a similar manner as in soymilk alone, but did not decrease the opaqueness of the solutions (as shown in FIG. 15), suggesting only soy components were precipitated while milk components remained soluble. A separate analysis of the weight of precipitated solids confirmed that the dry precipitated solids in skim milk and soymilk mixture is comparable to the amount of precipitated solids that would have precipitated from pure soymilk without skim milk (data not shown).

[0242] MgCh at concentrations higher than 0.4 M caused precipitation in bovine skim milk. In this experiment, MgCh was added in incremental doses to bovine skim milk while maintaining pH at 6.7 with 2 N NaOH. Magnesium chloride was added until the salt concentration reached 2.0 M. A 50 mL sample was taken at 0, 0.4, 0.8, 1.2, 1.6, and 2.0 M MgCh and labeled according to the salt concentrations. The samples were held at 50 °C for 20 minutes and centrifuged for 10 minutes at 3700 RPM. Casein began precipitating at 0.4 M MgCh and was nearly precipitated by 800 mM MgCh. This was evidenced by the reduction of turbidity and the consistent volume of the centrifuged pellet beyond 0.8 M, as shown in FIG. 16.

Example 6

[0243] In this example, the effect of adding glycinin or conglycinin on curd formation of milk proteins is compared. Skim milk is used as positive control. The procedure is described in FIGs 17-19. White Flake Soymilk (WFSM) is fractioned into a glycinin fraction and conglycinin fraction as shown in FIG. 17.

[0244] Soymilk Procedure

[0245] 1. Measure 100 g of dry ZFS Creston white flakes into a 1,000 mL beaker. Record actual weight and measured moisture.

[0246] 2 Add 900 mL water to a separate (empty) beaker. Heat water to 65 C. Targeting slurry total solids of 10%.

[0247] 3. Adjust pH to >8 with 2 N NaOH. Agitate on stir plate for 10 minutes.

[0248] 4. Centrifuge slurry for 10 minutes. Decant centrate from pellet. Save centrate and discard the okara fraction (pellet).

[0249] 5. Prior to use, centrifuge decanted soymilk for 5 minutes. Decant soymilk from pellet. Record volume/mass of centrate.

[0250] Glycinin Separation

[0251] 6. Add solid sodium bisulfate to achieve a concentration of 30 mM SO2. Adjust the pH to 6.0 with 2 N HCl.This should cause the glycinin fraction to “salt-out”.

[0252] 7. Centrifuge mixture for 10 minutes. Decant centrate from pellet.

[0253] 8. Combine pellets of glycinin and redisolve with water at ~10x the volume of the pellet. Adjust pH to 7.5 with 2 N NaOH. Record volume of pellet, water added, and final pH. [0254] 9. Set-up TFF System with a 100 kDa Synder PVDF membrane.

[0255] 10. Record volume and TS of starting material [0256] 11. Concentrate mixture to about 3x concentration factor (ex. 500 ml — > 167 mL) with the membrane pressure at 5 psig and room temperature.

[0257] 12. Dilute back to starting volume with water.

[0258] 13. Save mixture for cheese test. This is the glycinin fraction, which is enriched with glycinin and depleted in conglycinin.

[0259] Conglycinin Separation Adjust the pH of the supernatant of the last centrifugation step to 7.0 using 2 N NaOH.

[0260] 14. Prepare a solution (about 500 mL) of 20 mM Sodium Bisulfite.

[0261] 15. Set-up TFF System with a 100 kDa Synder PVDF membrane.

[0262] 16. Concentrate mixture to about 3x concentration factor (ex. 500 ml — > 167 mL) with the membrane pressure at 5 psig and room temperature.

[0263] 17. Dilute back to starting volume with water.

[0264] 18. Concentrate diluted mixture to a ~3x concentration factor with same membrane and filtration conditions.

[0265] 19. Save mixture for cheese test. This is the conglycinin fraction, which is enriched in conglycinin and depleted in glycinin.

[0266] The WFSM, glycinin fraction and conglycinin fraction were each subject to the process of making cheese as shown in FIG. 18 and FIG. 19.

[0267] Results: As shown in FIG. 20, Skim control (100% milk solids from skim milk) formed a curd as did 5% WFSM (White Flake Soymilk 5% weight + 95% weight milk solids). However, 15% WFSM (white flake soymilk solids 15% weight + 85% weight milk solids) did not form a curd. Both 5% glycinin (glycinin 5% weight + 95% weigh milk solids) and 15% glycinin (glycinin 15% weight + 85% weigh milk solids) formed curds.5% conglycinin (conglycinin 5% weight + 95% weigh milk solids) formed a loose curd, and 15% conglycinin (conglycinin 15% weight + 85% weigh milk solids) formed loose solids (i.e., poor coagulation). In all cases, the glycinin curds had at least one improved dairy characteristic as compared to conglycinin and WFSM blends. However, conglycinin enriched curds still had at least one improved dairy characteristic as compared to similar soy/milk ratios in WFSM which failed to form even loose curds at 15% soy solids.

[0268] FIG. 21 shows curd yield for different fractions. Replacing some casein protein with soy protein increased the curd protein yield compared to casein protein alone. A less expensive protein (soy) can replace a more expensive (dairy) protein and produce a cheese curd with higher yields on the input casein. At 5% weight (95% milk solids), glycinin has the highest curd dry solids/casein ratio, followed by conglycinin, while WSFM has the lowest curd dry solids/casein ratio. At 15% weight (85% milk solids), glycinin has the highest curd dry solids/casein ratio, followed by conglycinin which does not form solid curds, while WSFM failed to form any curd. [0269] FIG. 22 shows the total solids of the curds were measured for each curd produced in the experiment. The average total solids of curds made from glycinin/dairy blends were higher than conglycinin/dairy blends. A summary of data is shown in FIG. 23.

[0270] Elasticity of the curds was tested using the following procedure.

[0271] 1. Cheese plugs were cut from whole cheese curds using a 6.9mm cork borer

[0272] 2. Cheese plug dimensions were measured with calipers and recorded. Adjustments were made by cutting the cheese plugs with a razor blade if needed

[0273] 3. Plugs were placed flatly in glass test tubes

[0274] 4. The height of the cheese plugs in the test tubes were recorded

[0275] 5. A 10g weight was dropped onto the cheese in each test tube

[0276] 6. The depressed height of the cheese curd was recorded

[0277] 7. The weight was removed and the new height of the cheese in the test tube was recorded [0278] FIG. 27 shows curd elasticity measurements. Curd height, weighted height, and recovered height were measured for each curd. Glycinin curds had the closest elasticity to skim milk. Glycinin curds were firmer than the skim control, soymilk (also “WFSM”), and conglycinin curds. Skim milk curds had the largest change in height but recovered to nearly the same initial measurement. Conglycinin curds had poor elastic properties.

[0279] FIG. 28 shows Curd Elasticity % Return to Initial Height. The glycinin curds’ elasticity was closest to the control (skim) curd. The elasticity of the conglycinin and soymilk curds was reduced from the control.

[0280] Meltability of the curds was tested using the following procedure.

[0281] 1. Cheese disks were cut from whole curds with a 19.3mm metal whole punch

[0282] 2. Cheese disk dimensions were measured with calipers and recorded. Adjustments were made by cutting the cheese disks with a razor blade if needed

[0283] 3. IOOUL of vegetable oil was placed directly on the aluminum pan near the center.

Cheese disks were placed on top of the oil, and another IOOuL of vegetable oil was deposited on top of the cheese disk

[0284] 4. Once oiled, samples were placed in the oven at 90C for 5 minutes.

[0285] 5. Samples were allowed to cool for 30minutes at room temperature before remeasuring [0286] FIG. 29 shows cheese meltability. The observed stretchability of the curds containing soy was less than the skim control curd. The conglycinin’ s stretchability was impacted more so than the other soy containing curds. Example 7: In-vitro Micelle Formation (milk making from individual casein proteins)

[0287] - Dissolve 0.324g Potassium citrate tribasic in 1.5 ml water to get Tripotassium Citrate

[0288] - Dissolve 0.383g K2HPO4 in 11ml water to get Potassium phosphate.

[0289] - Dissolve 0470g CaC12-2H20 in 15 ml of water to get Calcium Chloride.

[0290] - Extract individual casein proteins from source organism or acquire purified caseins from sigma Aldrich.

[0291] Dissolve casein proteins in water to a concentration of 50 mg/ml to get casein water (or casein containing water).

[0292] Add ImL of casein containing water to a 5mL beaker

[0293] Add a mini stir bar

[0294] Place the 5mL beaker inside of a 600mL beaker on a hot plate containing approximately 125mL of water to submerge the 5mL beaker about a third of the height, using a clamping system (shown at the bottom of the page).

[0295] Make sure the 5mL beaker is not touching the bottom or the sides of the 600mL beaker.

[0296] Place a thermometer in the outer water with the bulb not touching the glass but fully submerged in the water

[0297] Set the hotplate around 65C after all of the casein is in solution adjust accordingly to make sure the water temperature stays at 37 °C.

[0298] Try to get the water temperature to 37 °C quickly to avoid too much evaporation.

[0299] Check the temperature on the thermometer often!

[0300] Set the stirring to 1000 RPM

[0301] Add the following

[0302] 20uL tripotassium citrate

[0303] 70uL potassium phosphate is

[0304] Wait 4 minutes.

[0305] Then every 4 minutes add (12x)

[0306] 12.5uL Potassium Phosphate Sol.

[0307] 25uL Calcium Chloride Sol.

[0308] Let stir with the temperature at 37 °C for 1 hour

[0309] Turn the heat off

[0310] Add 240uL water

[0311] Add 180uL Heavy Cream

[0312] The resulting composition will contain casein in micellar form.

Example 8: Cheese Making [0313] Any process for cheese making will be sufficient to make cheese from milk. Milk is heated in a large pot to 85-100°C and then cooled down to around 33-38°C. Lactic acid bacteria is added to the milk. Once the milk has reached the desired acidity level, rennet is added. The milk will coagulate and form curds. The curds are then cut into small pieces and heated again, which releases additional whey. The curds are kneaded and stretched until they form a smooth, elastic texture.

Example 9: Formation of In-vivo micelles

[0314] Vectors: A multigene vector, termed pMOZ1066, which expresses the coding regions 1) bovine a-Sl-casein (Uniprot accession # P02662) (SEQ ID No. 1), 2) green fluorescent protein (GFP, Uniprot accession # P42212) (SEQ ID No. 2), 3) bovine 0-casein (Uniprot accession # P02666) (SEQ ID No. 3), 4) bovine K-casein (Uniprot accession # P02668) (SEQ ID No. 4), and 5) bovine FAM20C kinase (uniprot accession number # F1MXQ3) (SEQ ID No. 5) was assembled using the modular cloning system MoClo (Engler, Carola, Mark Youles, Ramona Gruetzner, Tim-Martin Ehnert, Stefan Werner, Jonathan D. G. Jones, Nicola J. Patron, and Sylvestre Marillonnet. “A Golden Gate Modular Cloning Toolbox for Plants.” ACS Synthetic Biology 3, no. 11 (November 21, 2014): 839-43. https://doi.org/10.1021/sb4001504). All five proteins were expressed under constitutively active plant promoters, with a subset of these proteins possessing translationally-fused epitope tags and/or target-peptide sequences on their C- termini. Specifics on each gene subunit is outlined below.

[0315] Caseins-. Gene sequences that encode for bovine a-Sl-casein (P02662), bovine P-casein (P02666), and bovine K-casein (P02668) were derived from Uniprot and modified to include the following C-terminus 60bp sequence: (AGTTCG)(GATTACAAAGATGACGACGATAAG)(CATCATCACCATCACCAC)(CATGA TGAGTTG). The first sequence encodes for a two-serine spacer that functions to provide enough space for each casein protein to properly fold without steric hindrance or interference from other tags. The second and third sequences encode for two affinity purification tags (flag-tag and 6- Histidine tag) that are commonly used for protein identification and purification. The fourth sequence encodes for an HDEL target peptide that functions to retain soluble casein proteins in the endoplasmic reticulum (ER). Additionally, an N-terminus signal peptide, gmGlycininl (GY1, P04776), was added to all three caseins in order to target them towards the ER and vacuoles. Recombinant casein sequences were expressed under the constitutive AtuMas promoter and 5’ untranslated region (UTR) (https://www.ncbi.nlm.nih.gov/pmc/articles/PMC287101/).

[0316] Gene sequence encoding for bovine K-casein (P02668) was derived from Uniprot and modified to include the following 57bp sequence to increase the hydrophilicity of the C-terminus region, following the sequence (SEQ ID No. 7): TCGGATTACAAAGATGACGACGATAAGCATCATCACCATCACCACCATGATGAGTTG , corresponding to amino acid sequence SDYKDDDDKHHHHHHHDEL (SEQ ID No. 8) [0317] Additionally, an N-terminus signal peptide, gmGlycininl (GY1, P04776), was added to all three caseins in order to target them towards the ER and vacuoles. Recombinant casein sequences were expressed under the constitutive AtuMas promoter and 5’ untranslated region (UTR).

[0318] FAM20C kinase: The gene sequence of for the FAM20C kinase (F1MXQ3) was modified to include the N-terminus At5g67360 signal peptide sequence and a C-terminus HDEL target peptide sequence (CACGATGAGTTA). The recombinant FAM20C kinase was expressed under the 35S short promoter (https://www.ncbi.nlm.nih.gov/pmc/articles/pmid/16578811/).

[0319] Green fluorescent protein (GFP)'. Cytosolic GFP (P42212) was expressed under the 35S short promoter and used as a visual reporter for detecting the expression of pMOZ1066 in transient soybean agrobacterium transformations.

[0320] E.coli transformation'. pMOZ1066 (FIG. 25, SEQ ID NO. 9) was transformed into Lucigen Ecloni 10G bacteria using the following chemical transformation protocol: First, Lucigen Ecloni 10G E.coli thermo-competent cells were thawed on ice for approximately 5 minutes. Then, competent cells were spiked with 1 pg-100 ng of plasmid DNA and left to incubate on ice for 10 minutes. Once complete, the plasmid-bacterial mixture was heat shocked at 42°C for 90 seconds and immediately replaced on ice for 5 minutes. Afterwards, transformed cells were mixed with lOOpl of liquid broth (LB) and cultured in a 37°C shaker set to 225 rpm for 45 minutes. Following incubation, cultured bacteria was plated on LB agar plates with the appropriate antibiotic for selection (1 : 1000 concentration) and left to further incubate overnight in a 37°C growth chamber.

[0321] High throughput blue-white selection'. Following overnight incubation, plates were checked for bacterial colony growth. To increase recombinant bacteria selection, a MoClo compatible blue-white selection system was used. In brief, blue-white selection plasmid vectors carry the “lacZ” operon sequence within their multiple cloning site. In the absence of recombinant DNA, the lacZ operon will enable a biochemical reaction that turns the colony blue. Whereas, when recombination occurs, lacZ operon activity will be disrupted leaving the colony to be white. For each cloning reaction, at least two white colonies were chosen for further verification.

[0322] Plasmid selection and verification: Picked colonies were placed in 5 mLs of LB plus their respective selection antibiotic (1 : 1000 concentration) and cultured overnight in a 37°C shaker (225 rpm). Once cloudy, plasmids were purified out of bacteria using the NucleoSpin miniprep kit (Takara bio inc.). Then, each plasmid was digested using the appropriate restriction enzymes in order to confirm the presence of a DNA insert and sent for sanger and nanopore sequencing to confirm sequence correctness. Colonies containing the correct plasmids were made into frozen 20% glycerol stocks.

[0323] Electroporation into agrobacterium'. The completed pMOZ1066 plasmid was transformed into EHA105 electrocompetent agrobacterium cells. To achieve this, 30ng of pMOZ1066 purified plasmid was mixed into ice-thawed electrocompetent cells and swiftly transferred into a pre-chilled 0.2 cm Gene pulser cuvette. The cuvette was then loaded into an electroporation chamber and given an electric pulse of 2.5kV. The resulting transformed cells were mixed with 500 pl of LB and left to culture in a 28°C shaking incubator (120rpm) for 2-4 hours. Afterwards, cultured cells were plated onto LB agar plates containing the appropriate antibiotic selection media and placed in a 28°C incubator for two days. Once colonies formed, a minimum of three were picked for further verification via purification, digestion, and sequencing. [0324] Transient agrobacterium transformation into zygotic soy embryos: Zygotic soybeans were transiently transformed using agrobacterium. Specifically, a 5mL starter culture of pMOZ1066 agrobacterium was started from either a glycerol stock or bacterial colony and incubated for two days in a 28°C shaker (120rpm). Once cloudy, the 5mL starter culture was used to inoculate a larger 200mL overnight culture.

[0325] Bacteria preparation '. Large pMOZ1066 agrobacterium cultures were spun down in a large centrifuge at 3400g for 10 minutes. The supernatant was then removed and the remaining agrobacterium pellet was resuspended in 30mL of LCCM. Post resuspension, the agrobacterium was centrifuged at 3400g for 6 minutes, and then subjected to one more round of supernatant removal, pellet resuspension and centrifugation. After the final spindown, the remaining supernatant was removed and the bacterial pellet was resuspended in lOmL of LCCM. An OD600 measurement was then taken and the agrobacterium solution was diluted to a final concentration of 1.4 OD600. The resulting agrobacterium solution was spiked with fresh acetosyringone (lOOpM final concentration) and subsequently incubated at room temperature while shaking (120rpm) for 1-2 hours.

[0326] Seed sterilization and preparation: During the agrobacterium incubation period, pods were picked from soy plants containing 8-10mm embryos (~8 weeks old) and sterilized by the following method: 70% ethanol bath for 30 seconds, 10% bleach bath for 10 minutes, three sequential sterilized deionized water bath for 5 minutes each. After sterilization, seeds were aseptically removed from their pods, dissected from their seed coats, and split in half.

[0327] Explant inoculation'. Once agrobacterium cultures finished incubating, silwet-77 (0.03% v/v) was added and mixed until dissolved. Then, 30 cotyledon halves (15 explants) were placed in the agrobacterium culture, and sonicated for 20 seconds at a 20% amplitude with 5s/l/s on/off pulse cycles. After sonication, explants were vacuum infiltrated for 5 minutes and left to incubate for two hours on a room temperature rotator.

[0328] Plating explants-. Explants were removed from bacterial culture and placed flat down (adaxial side down) on SCCM plates with a layer of sterile filter paper. Plates were then wrapped with micropore tape and incubated for 3 days in a dark 24°C chamber.

[0329] Washing explants: After plants incubated in the dark for 3 days, they were washed three times for five minutes each with sterile water containing Rif, Carb+Cef. Then, they were replated on SCCM plates containing filter paper and left to incubate in a dark 24°C chamber for another 7-9 days.

[0330] Casein purification and characterization:

[0331] Protein Crude Extraction: Roughly 40 plates containing pMOZ1066 transformed cotyledons were flash frozen in liquid nitrogen and crushed into a fine powder. Then, 100 grams of powder was measured out and mixed with a tris protein extraction buffer (50 mM Tris, 300 mM KC1, 0.5% Tween-20, 3.65% glycerol, Sigma plant protease inhibitor, pH 8.6). The resulting mixture rotated for 1 hour at 4°C and then spundown at 1300rpm for 30 minutes at 4°C.

[0332] Casein purification: Crude protein extract was first clarified using a ,45pm filter. Then, the sample was mixed with nickel resin, rotated at 4°C for 4 hours, and subsequently centrifuged for 2 minutes at 1000g. After centrifugation, the supernatant was removed and the remaining sample was washed with a wash buffer (50mM Tris base, 300mM KCL, 20mM imidazole, pH 7.4). To remove impurities, the wash step was repeated four times. After the last wash, the proteins were rotated in an elution buffer at 4°C for 15 minutes. Then, they were centrifuged at 700g for two minutes. The supernatant was saved for later analysis and the elution step was repeated another four times.

[0333] Casein characterization:

[0334] To confirm successful purification of the caseins, the first three elution samples were run on a BioRad TGX AnykD Mini Protean SDS-PAGE gel and blotted with antibodies against FLAG to check for the presence of the caseins. “WT” is wildtype control. “Flag” ispositive control for flag tag. “WT’ is wash l. “El” (elution 1), “E2” (elution 2) and “E3” (elution 3) are serial elutions. This blot was compared to the initial flow through and wash supernatants, which were expected to have minimal FLAG detection. Positive bands were enriched in the elution samples and present at the expected size of 27kDa. As all three caseins have Flag tag, mass spectrometry was used to confirm the presence of all three proteins.

[0335] Immunogold electron microscopy for micelle validation'. Casein micelles have been previously shown to be detectable using transmission electron microscopy (TEM). Therefore, recombinant milk, synthetic milk, non-fat raw bovine milk, and a purified sample of non-casein soy proteins, were all fixed in a 0.1% PFA solution, embedded in LR white resin, and immunolabeled with commercially available beta polyclonal antibodies (ProSci did the rabbits, Pacific Immunology did the goats, Bioss antibodies) and colloidal gold-conjugated 6 nm secondary antibodies (Ted Pella, inc. or Electron Microscopy Sciences). Each sample was imaged on a Talos L120C G2 transmission electron microscope using practices known to those skilled in the art. Each sample possessed spherical protein-assembled structures but only the samples containing caseins were decorated with anti -beta-bound gold particles (shown in FIG. 26).

Table 1. Sequences IDs used in Example 9.

Prophetic Example A

[0336] In this proposed example, plant material is defatted in a hexane extraction process and subsequently processed according to salt coagulation and membrane filtration process described in Example la. In soybeans, the fat extracted in the soymilk process is removed with the coagulated protein. It is expected that removing the fat with the plant material will minimize membrane fouling from fat/oil. Plant oils are high in value. If the process economics dictate the fat first be captured, it is expected that the described process could still isolate and purify the recombinant protein.

Prophetic Example B

[0337] In this proposed example a whole soymilk is first supplemented with dairy skim milk. The native plant proteins are coagulated and precipitated with magnesium chloride. The precipitated solids are removed in a centrifuge step and the supernatant collected. The ionic strength of the supernatant will be adjusted with sodium chloride to an ionic strength of 0.2 M. The salty supernatant is then concentrated and diafiltered with water containing sodium chloride at ionic strengths ~0.2 M. It is expected that this example will improve the purity of the final retentate containing the casein protein.

[0338] It should be understood that the description and the drawings are not intended to limit the invention to the particular form disclosed, but to the contrary, the intention is to cover all modifications, equivalents, and alternatives falling within the spirit and scope of the present invention as defined by the appended claims. Further modifications and alternative embodiments of various aspects of the invention will be apparent to those skilled in the art in view of this description. Accordingly, this description and the drawings are to be construed as illustrative only and are for the purpose of teaching those skilled in the art the general manner of carrying out the invention. It is to be understood that the forms of the invention shown and described herein are to be taken as examples of embodiments. Elements and materials may be substituted for those illustrated and described herein, parts and processes may be reversed or omitted, and certain features of the invention may be utilized independently, all as would be apparent to one skilled in the art after having the benefit of this description of the invention. Changes may be made in the elements described herein without departing from the spirit and scope of the invention as described in the following claims. Headings used herein are for organizational purposes only and are not meant to be used to limit the scope of the description.

Claims

WHAT IS CLAIMED IS:

1. A composition, comprising: casein micelles; and a soy ingredient comprising at least one of a 7s or a 1 Is, wherein the ratio of the 7s and the I ls (7s/l Is) is lower than a naturally occurring ratio.

2. The composition of claim 1, wherein the casein micelle comprises a recombinant casein protein.

3. The composition of claim 1 or 2, wherein the weight ratio of the 7s and the I ls (7s/l Is) is lower than a naturally occurring weight ratio, and the naturally occurring weight ratio of the 7s and the I ls (7s/l Is) is between 0.5-1.3.

4. The composition of claim 1 or 2, wherein the ratio of the 7s and the I ls (7s/l Is) is a weight ratio.

5. The composition of claim 1 or 2, wherein the ratio of the 7s and the I ls (7s/l Is) is a molar ratio.

6. The composition of any one of claims 1-5, wherein the ratio of the 7s and the I ls (7s/l Is) is lower than 0.5.

7. The composition of any one of claims 1-5, wherein the ratio of the 7s and the I ls (7s/l Is) is lower than 0.4.

8. The composition of any one of claims 1-5, wherein the ratio of the 7s and the I ls (7s/l Is) is lower than 0.3.

9. The composition of any one of claims 1-5, wherein the ratio of the 7s and the I ls (7s/l Is) is lower than 0.2.

10. The composition of any one of claims 1-5, wherein the ratio of the 7s and the I ls (7s/l Is) is lower than 0.1.

11. The composition of any one of claims 1-5, wherein the ratio of the 7s and the I ls (7s/l Is) is 0, wherein the soy ingredient does not comprise 7s.

12. The composition of any one of claims 1-5, wherein the ratio of the 7s and the I ls (7s/l Is) between 0 and 0.01, wherein the soy ingredient comprises a trace amount of the 7s, or does not comprise a detectable amount of the 7s.

13. The composition of any one of claims 1-12, wherein the soy ingredient is modified to decrease the amount of the 7s in the soy ingredient.

14. The composition of any one of claims 1-13, wherein the soy ingredient is modified to increase the amount of the 1 Is in the soy ingredient. The composition of any one of claims 1-14, wherein the soy ingredient is in the form of soy protein isolate. The composition of any one of claims 1-14, wherein the soy ingredient is in the form of soymilk. The composition of any one of claims 1-14, wherein the source of the casein micelles is from an animal. The composition of claim 17, wherein the animal is a mammal. The composition of claim 18, wherein the mammal is a ruminant. The composition of claim 19, wherein the ruminant is bovine. The composition of any of claims 1-20, wherein the source of the casein micelles is from a genetically modified microorganism. The composition of claim 21, wherein microorganism is at least one of yeast, fungi, or bacteria. The composition of any of claims 1-20, wherein the source of the casein micelles is from a genetically modified plant. The composition of any of claims 1-23, wherein casein protein is less than 50% (w/w) of total protein in the composition, or wherein casein protein is less than 60% (w/w) of total protein in the composition. The composition of any of claims 1-23, wherein casein protein is less than 70% (w/w) of total protein in the composition. The composition of any of claims 1-23, wherein casein protein is less than 80% (w/w) of total protein in the composition. The composition of any of claims 1-23, wherein casein protein is less than 90% (w/w) of total protein in the composition. The composition of any of claims 1-23, wherein casein protein is less than 100% (w/w) of total protein in the composition. The composition of any of claims 1-28, wherein soy protein is more than 30% (w/w) of total protein in the composition. The composition of any of claims 1-28, wherein soy protein is more than 40% (w/w) of total protein in the composition. The composition of any of claims 1-28, wherein soy protein is more than 50% (w/w) of total protein in the composition. The composition of any of claims 1-28, wherein soy protein is more than 60% (w/w) of total protein in the composition. The composition of any of claims 1-28, wherein soy protein is more than 70% (w/w) of total protein in the composition. The composition of any of claims 1-28, wherein soy protein is more than 80% (w/w) of total protein in the composition. The composition of any of claims 1-28, wherein soy protein is more than 90% (w/w) of total protein in the composition. The composition of any one of claims 1-35, wherein the composition forms a cheese curd in a cheese making process. The composition of claim 36, wherein the composition forms a firm cheese curd in a cheese making process. The composition of claim 37, wherein the composition forms a firm cheese curd in a cheese making process within 15 minutes in the cheese making process. The composition of claim 37, wherein the composition forms a firm cheese curd in a cheese making process within 30 minutes in the cheese making process. The composition of claim 37, wherein the composition forms a firm cheese curd in a cheese making process within 60 minutes in the cheese making process. The composition of claim 37, wherein the composition forms a firm cheese curd in a cheese making process within 100 minutes in the cheese making process. The composition of claim 36, wherein the composition forms a soft cheese curd in a cheese making process. The composition of claim 42, wherein the composition forms a soft cheese curd within 15 minutes in the cheese making process. The composition of claim 42, wherein the composition forms a soft cheese curd within 30 minutes in the cheese making process. The composition of claim 42, wherein the composition forms a soft cheese curd within 60 minutes in the cheese making process. The composition of claim 42, wherein the composition forms a soft cheese curd within 100 minutes in the cheese making process. The composition of any one of claims 1-46, wherein the cheese making process is performed at pH between 6.0-7.0. The composition of any one of claims 1-46, wherein the cheese making process is performed at pH between 6.0-6.5. The composition of any one of claims 1-46, wherein the cheese making process is performed at pH between 6.5-7.0. The composition of any one of claims 36-49, wherein the cheese making process comprises adding rennet to the composition to cause the casein micelle to precipitate. The composition of any one of claims 1-50, wherein the soy ingredient comprises a non- detectable or reduced level of an anti -nutritional or toxin in soybean, compared to a naturally occurring level. The composition in claim 51, wherein the anti -nutritional or toxin comprises at least one of trypsin inhibitor, phytic acid, lectins, or soybean toxin. The composition of any one of claims 1-52, wherein the 1 Is is glycinin. The composition of any one of claims 1-53, wherein the 7s is P-conglycinin. A dairy product or dairy product substitute comprising the composition in any one of claims 1-54. The dairy product or dairy product substitute in claim 55, wherein the dairy product or dairy product substitute is cheese, cheese curd, or milk. A method of making cheese, comprising, in sequence, providing the composition in any one of claims 1-54; and adding an enzyme to the composition to cause the casein micelles to precipitate. The method in claim 57, further comprising mixing a soy ingredient with the casein micelles. The method in claim 57, wherein the enzyme is added after mixing soy ingredient with the casein micelles. The method in any one of claims 57-59, wherein the enzyme comprises at least one enzyme found in rennet. The method in any one of claims 57-59, wherein the enzyme comprises a protease. The method in any one of claims 57-59, wherein the enzyme comprises at least one of chymosin, pepsin or lipase. A method of making a dairy product, comprising: providing a liquid mixture comprising casein micelles and at least one soy protein; removing a portion of the soy protein from the liquid mixture; and adding an enzyme to the liquid mixture to cause the casein micelles to precipitate. The method in claim 63, wherein at least one soy protein soy protein comprises a subunit of conglycinin. The method in claim 64, wherein the subunit is a subunit or a’ subunit of conglycinin. The method in any one of claims 63-65, wherein removing the portion of the soy protein from the liquid mixture decreases the ratio of the conglycinin to the glycinin (conglycinin/glycinin). The method in any one of claims 63-66, wherein the enzyme comprises at least one enzyme found in rennet. The method in any one of claims 63-66, wherein the enzyme comprises a protease. The method in claim 68, wherein the enzyme comprises at least one of chymosin, pepsin or lipase. The method in any one of claims 63-69, wherein removing the portion of soy protein from the mixture comprises adding a salt to the liquid mixture to precipitate the portion of soy protein. The method in claim 70, wherein the salt is at least one of sodium phosphate, calcium chloride, or potassium chloride. The method in any one of claims 63-71, wherein removing the portion of soy protein from the mixture further comprises filtering the composition to produce a supernatant. The method in claim 72, wherein the supernatant has less of the portion of the soy protein that does the liquid mixture before filtering. The method in claim 72, wherein the supernatant has a lower concentration of conglycinin than does the liquid mixture before filtering. The method in claim 72, wherein the supernatant has a lower ratio of conglycinin to glycinin (conglycinin/glycinin) does the liquid mixture before filtering. The method in claim 72, wherein filtering the composition comprising using a microfiltration (MF) membrane to filter the composition. The method in claim 76, wherein filtering the composition comprising using a benchtop tangential flow filtration system to filter the composition. The method in claim 72, wherein the microfiltration (MF) membrane has a pore size between 0.1 and 10 pm. The method in claim 72, wherein the microfiltration (MF) membrane has a pore size between 0.1 and 1 pm. The method in claim 72, wherein the microfiltration (MF) membrane has a pore size between 1 and 10 pm. The method in claim 72, wherein the microfiltration (MF) membrane has a pore size smaller than 0.1 pm. The method in any one of claims 63-80, wherein the casein micelles precipitate to form a curd. The method in any one of claims 63-80, wherein the casein micelles precipitate to form a solid or semi-solid. A dairy product or dairy product substitute made using the methods in any one of claims 51-

83. The dairy product or dairy product substitute in claim 84, wherein the dairy product or dairy product substitute is cheese. A composition, comprising: a casein micelle derived from a plant; and a plant protein comprising one of legumin, vicilin, prolamin, gliadin, 0-conglycinin, or glycinin, or any combination thereof. The composition in claim 86, wherein the plant protein comprises I ls. The composition in claim 87, wherein the plant protein further comprises 7s. The composition of claim 88, wherein the ratio of 7s/l Is is lower than a naturally occurring ratio. The composition of claim 89, wherein the naturally occurring ratio is between 0.5 and 1.3. The composition of claim 86, wherein the casein micelle is formed in vivo in a plant. The composition of claim 86, wherein the casein micelle is formed in vitro. The composition of claim 86, wherein the casein micelle comprises bovine casein. The composition of claim 86, wherein the casein micelle comprises human casein. The composition of claim 88, wherein the ratio of 7s/l Is (w/w) is less than 0.4. The composition of claim 88, wherein the ratio of 7s/l Is (w/w) is less than 0.3. The composition of claim 88, wherein the ratio of 7s/l Is (w/w) is less than 0.2. The composition of claim 88, wherein the ratio of 7s/l Is (w/w) is less than 0.1. The composition of claim 86 or 87, wherein the composition does not comprise a detectable amount of 7s. . The composition in any one of claims 86-99, wherein the plant protein is inside the casein micelle. . The composition in any one of claims 86-100, wherein an outer layer of the casein micelle is enriched with a K-casein, and an inner matrix of the casein micelle comprises at least one of aS 1 -casein, aS2-casein, or 0- casein. . The composition in claim 101, wherein the plant protein interacts with one or more of the casein proteins in the inner matrix. . The composition in claim 102, wherein the plant protein binds to any one of aS 1 -casein, aS2-casein, or 0-casein. . The composition in any one of claims 101-103, wherein the plant protein is at least partially embedded in the inner matrix.

. The composition in claim 104, wherein the plant protein is fully embedded in the inner matrix. . The composition in any one of claims 86-99, wherein the plant protein is on the surface of the casein micelle. . The composition in claim 105, wherein the plant protein interacts with a K-casein on an outer layer of the casein micelle. . The composition in claim 107, wherein the plant protein binds to the K-casein. . The composition in any one of claims 106-108, wherein the plant protein is at least partially embedded in the outer layer of the casein micelle. . The composition in any one of claims 86-108, further comprising a plant lipid, a plant sugar, a plant polyphenol, or plant DNA molecule, or any combination thereof. . The composition in claim 110, wherein the plant lipid, a plant sugar, a plant polyphenol, or plant DNA molecule is in a trace amount. . The composition in claim 110, wherein the plant lipoid comprises at least one of stigmasterol, sitosterol, campesterol, or brassicasterol. . The composition in claim 110, wherein the plant polyphenol comprises isoflavone. . The composition in claim 110, wherein the plant DNA comprises plant DNA fragments.. The composition in any one of claims 86-114, wherein the plant protein is in a trace amount.. The composition in claim 115, wherein the plant protein is less than 5% (w/w) of total protein content in the composition. . The composition in claim 115, wherein the plant protein is less than 3% (w/w) of total protein content in the composition. . The composition in claim 115, wherein the plant protein is less than 1% (w/w) of total protein content in the composition. . The composition in claim 115, wherein the plant protein is less than 0.5% (w/w) of total protein content in the composition. . The composition in claim 115, wherein the plant protein is less than 0.1% (w/w) of total protein content in the composition. . The composition in claim 115, wherein the plant protein is less than 0.05% (w/w) of total protein content in the composition. . The composition in claim 115, wherein the plant protein is less than 0.01% (w/w) of total protein content in the composition. . The composition in any one of claims 115-122, wherein the plant protein is more than 0% (w/w) of total protein content in the composition.

. The composition in any one of claims 115-121, wherein the plant protein is more than 0.005% (w/w) of total protein content in the composition. . The composition in any one of claims 115-120, wherein the plant protein is more than 0.01% (w/w) of total protein content in the composition. . The composition in any one of claims 115-119, wherein the plant protein is more than 0.05% (w/w) of total protein content in the composition. . The composition in any one of claims 115-118, wherein the plant protein is more than 0.1% (w/w) of total protein content in the composition. . The composition in any one of claims 115-117, wherein the plant protein is more than 0.5% (w/w) of total protein content in the composition. . The composition in any one of claims 115-116, wherein the plant protein is more than 1% (w/w) of total protein content in the composition. . The composition in any one of claims 115-116, wherein the plant protein is more than 3% (w/w), or more than 5% (w/w) of total protein content in the composition. . The composition in any one of claims 86-130, wherein the composition does not comprise a detectable amount of a-lactalbumin. . The composition in any one of claims 86-131, wherein the composition does not comprise a detectable amount of P-lactoglobulin. . The composition in any one of claims 86-132, wherein the composition does not comprise a detectable amount of a-S2-casein. . The composition in any one of claims 86-133, wherein the composition does not comprise a detectable amount of lactoferrin. . The composition in any one of claims 86-134, wherein the composition does not comprise a detectable amount of transferrin. . The composition in any one of claims 86-135, wherein the composition does not comprise a detectable amount of serum albumin. . The composition in any one of claims 86-136, wherein the composition does not comprise a detectable amount of lysozyme. . The composition in any one of claims 86-137, wherein the composition does not comprise a detectable amount of lactoperoxidase. . The composition in any one of claims 86-138, wherein the composition does not comprise a detectable amount of immunoglobulin-A. . The composition in any one of claims 86-139, wherein the composition does not comprise a detectable amount of lipase. . The composition in any one of claims 86-140, wherein the composition is a powder.

. The composition in any one of claims 86-141, further comprising a coagulant. . The composition in claim 142, wherein the coagulant is in a detectable amount. . The composition in claim 143, wherein the coagulant is a magnesium salt. . The composition in claim 144, wherein the magnesium salt is at least one of magnesium chloride (MgC12) or magnesium sulfate (MgSO4). . The composition in claim 142 or 143, wherein the coagulant is a calcium salt. . The composition in claim 146, wherein the calcium salt is at least one of calcium chloride (CaC12) or calcium sulfate (CaSO4). . The composition in claim 142 or 143, wherein the coagulant is glucono-delta-lactone (GDL). . The composition in any one of claims 142-148, wherein the coagulant is less than 0.1% (w/w) the composition. . The composition in any one of claims 142-148, wherein the coagulant is less than 0.05% (w/w) the composition. . The composition in any one of claims 142-148, wherein the coagulant is less than 0.01% (w/w) the composition. . The composition in any one of claims 142-148, wherein the coagulant is less than 0.005% (w/w) the composition. . The composition in any one of claims 142-148, wherein the coagulant is less than 0.001% (w/w) the composition. . The composition in any one of claims 142-148, wherein the coagulant is more than 0.0005% (w/w) the composition. . The composition in any one of claims 142-148, wherein the coagulant is more than 0.0001% (w/w) the composition. . The composition in any one of claims 142-155, wherein the composition further comprises a fat. . The composition in claim 156, wherein the composition has a total fat content below 1%.. The composition in claim 156, wherein the composition has a total fat content below 0.1%.. The composition in claim 156, wherein the composition has a total fat content below 0.01%.. The composition in claim 156, wherein the composition does not comprise a detectable amount of animal fat. . The composition in any one of claims 86-160, wherein the composition does not comprise a detectable amount of lactose. . The composition in any one of claims 86-161, wherein the composition does not comprise a detectable amount of animal hormone.

. The composition in any one of claims 86-161, wherein the composition does not comprise a detectable amount of estrogen. . The composition in any one of claims 86-163, wherein the composition does not comprise a detectable amount of progesterone. . The composition in any one of claims 86-164, wherein the composition does not comprise a detectable amount of corticoid. . The composition in any one of claims 86-165, wherein the composition does not comprise a detectable amount of androgen. . The composition in any one of claims 86-166, wherein the composition is a substitute dairy product. . The composition in any one of claims 86-167, wherein the casein micelles derived from plants confer on the composition one or more characteristics of a dairy product selected from the group consisting of: taste, flavor, aroma, appearance, mouthfeel, density, structure, texture, elasticity, springiness, coagulation, binding, leavening, aeration, foaming, creaminess, and emulsification. . The composition in claim 168, wherein the dairy product is any one of milk, milk powder, cheese, yogurt, ghee, or butter. . The composition in any one of claims 86-169, wherein the composition is homogenous.. The composition in any one of claims 86-169, wherein the composition is substantially homogenous. . The composition in claim 115, wherein the plant protein is less than 0.005% (w/w) of total protein content in the composition. . The composition in claim 115, wherein the plant protein is less than 0.0001% (w/w) of total protein content in the composition. . The composition in claim 115, wherein the plant protein is less than 0.00005% (w/w) of total protein content in the composition. . The composition in any one of claims 86-174, wherein the plant protein is more than 0.00001% (w/w) of total protein content in the composition. . The composition in any one of claims 86-174, wherein the plant protein is more than 0.00003% (w/w) of total protein content in the composition. . The composition in any one of claims 86-173, wherein the plant protein is more than 0.00005% (w/w) of total protein content in the composition. . The composition in any one of claims 86-172, wherein the plant protein is more than 0.0001% (w/w) of total protein content in the composition.

. The composition in any one of claims 86-114, wherein the plant protein is more than 5% (w/w) of total protein content in the composition. . The composition in any one of claims 86-114, wherein the plant protein is more than 10% (w/w) of total protein content in the composition. . The composition in any one of claims 86-114, wherein the plant protein is more than 20% (w/w) of total protein content in the composition. . The composition in any one of claims 86-114, wherein the plant protein is more than 30% (w/w) of total protein content in the composition. . The composition in any one of claims 86-114, wherein the plant protein is more than 40% (w/w) of total protein content in the composition. . The composition in any one of claims 86-114, wherein the plant protein is more than 50% (w/w) of total protein content in the composition. . The composition in any one of claims 86-114, wherein the plant protein is more than 60% (w/w) of total protein content in the composition. . The composition in any one of claims 86-114, wherein the plant protein is more than 70% (w/w) of total protein content in the composition. . The composition in any one of claims 86-114, wherein the plant protein is more than 80% (w/w) of total protein content in the composition. . The composition in any one of claims 86-179, wherein the plant protein is less than 10% (w/w) of total protein content in the composition. . The composition in any one of claims 86-180, wherein the plant protein is less than 20% (w/w) of total protein content in the composition. . The composition in any one of claims 86-181, wherein the plant protein is less than 30% (w/w) of total protein content in the composition. . The composition in any one of claims 86-182, wherein the plant protein is less than 40% (w/w) of total protein content in the composition. . The composition in any one of claims 86-183, wherein the plant protein is less than 50% (w/w) of total protein content in the composition. . The composition in any one of claims 86-184, wherein the plant protein is less than 60% (w/w) of total protein content in the composition. . The composition in any one of claims 86-185, wherein the plant protein is less than 70% (w/w) of total protein content in the composition. . The composition in any one of claims 86-186, wherein the plant protein is less than 80% (w/w) of total protein content in the composition.

. The composition in any one of claims 86-187, wherein the plant protein is less than 90% (w/w) of total protein content in the composition. . A composition comprising a casein micelle and a soy ingredient, wherein the soy ingredient comprises 1 Is in an amount greater than 90% (w/w) of total soy proteins. . A composition comprising a casein micelle and a soy ingredient, wherein the soy ingredient comprises 1 Is in an amount greater than 80% (w/w) of total soy proteins. . A composition comprising a casein micelle and a soy ingredient, wherein the soy ingredient comprises 1 Is in an amount greater than 70% (w/w) of total soy proteins. . A composition comprising a casein micelle and a soy ingredient, wherein the soy ingredient comprises 1 Is in an amount greater than 60% (w/w) of total soy proteins. . A composition comprising a casein micelle and a soy ingredient, wherein the soy ingredient comprises 1 Is in an amount greater than 50% (w/w) of total soy proteins. . The composition in any one of claims 86-201, wherein the composition is a dairy product or a dairy product substitute. . The composition in any one of claims 86-201, wherein the composition is an intermediate product made during making a dairy product. . The composition in any one of claims 86-201, wherein the composition is a cheese product.. A method of extracting a recombinant protein expressed in a plant, comprising: obtaining a mixture comprising water, the recombinant protein, and a component from the plant; adding a coagulant to the mixture to cause at least a portion of the plant component to coagulant; and extracting supernatant from the mixture, wherein the supernatant comprises the recombinant protein. . The method in claim 205, wherein the plant is at least one of alfalfa, cassava, cotton, cowpea, maize, pea, peanut, rice, sesame, sorghum, soybean, or yam. . The method in claim 205 or 206, the methods are performed in sequence. . The method in any one of claim 205-207, wherein the mixture comprises a slurry made by blending the plant in water. . The method in any one of claim 205-208, further comprising separating insoluble plant components from the slurry to produce a plant-based milk before adding the coagulant to the mixture. . The method in any one of claim 205-209, further comprising adjusting pH of the mixture after adding the coagulant.

. The method in claim 210, further comprising adding additional coagulant after adjusting pH of the mixture. . The method in claim 210, wherein the additional coagulant is different from the coagulant. The method in claim 210, further comprising adjusting pH of the mixture after adding the additional coagulant. . The method in any one of claims 205-213, wherein the recombinant protein is a casein protein. . The method in any one of claims 205-213, wherein the recombinant protein is a whey protein. . The method in any one of claims 205-213, wherein the recombinant protein is an egg white protein. . The method in any one of claims 205-213, wherein the recombinant protein is a collagen protein. . The method in any one of claims 205-217, wherein the coagulant is at least one of a magnesium salt, a calcium salt, glucono-delta-1 actone (GDL), or any combination thereof.. The method in claim 218, wherein the calcium salt is at least one of calcium chloride (CaCh) or calcium sulfate (CaSC ). . The method in claim 218, wherein the magnesium salt is at least one of magnesium chloride (MgCh) or magnesium sulfate (MgSC ). . The method in any one of claims 1-220, wherein the coagulant is added to the mixture reach a concentration between 20 and 400 mM. . The method in claim 221, wherein the coagulant is added to the mixture reach a concentration between 40 and 400 mM. . The method in claim 221, wherein the coagulant is added to the mixture reach a concentration between 20 and 300 mM. . The method in claim 221, wherein the coagulant is added to the mixture reach a concentration between 40 and 300 mM. . The method in claim 221, wherein the coagulant is added to the mixture reach a concentration between 40 and 200 mM. . The method in claim 221, wherein the coagulant is added to the mixture reach a concentration between 20 and 200 mM. . The method in any one of claims 205-226, further comprising at least one of: cleaning the plant material to remove dirt and foreign material; dehulling or deshelling the plant material; flaking the plant material; reducing the particle size of the plant material; extracting oil from the plant material with a hexane based solvent; desolventizing the plant material without cooking and denaturing the recombinant protein; soaking the plant material in water; or any combination thereof. . The method in any one of claims 205-227, wherein the method is performed below 80 °C.. The method in any one of claims 228, wherein the method is performed at ambient temperature. . The method in any one of claims 228, wherein the method is performed between 0 °C and 80 °C. . The method in any one of claims 228, wherein the method is performed between 0 °C and 70 °C. . The method in any one of claims 228, wherein the method is performed between 0 °C and 60 °C. . The method in any one of claims 228, wherein the method is performed between 0 °C and 50 °C. . The method in any one of claims 228, wherein the method is performed between 0 °C and 40 °C. . The method in any one of claims 228, wherein the method is performed between 0 °C and 30 °C. . The method in any one of claims 228, wherein the method is performed between 10 °C and 25 °C. . The method in any one of claims 205-236, further comprising filtering the supernatant.. The method in claim 237, wherein filtering the supernatant comprising using a microfiltration (MF) membrane. . The method in claim 238, wherein the microfiltration (MF) membrane has a pore size between 0.1 and 10 pm. . The method in claim 238, wherein the microfiltration (MF) membrane has a pore size between 0.1 and 1 pm. . The method in claim 238, wherein the microfiltration (MF) membrane has a pore size between 1 and 10 pm. . The method in claim 238, wherein filtering the supernatant at least partially remove a plant component include soluble proteins, sugars, and minerals.

. The method in any one of claims 205-236, further comprising a diafiltration step to purify the protein. . The method of claim 243, wherein the diafiltration step uses a solution comprising at least one of magnesium chloride, sodium chloride, or calcium chloride. . A dairy product or dairy product substitute comprising the supernatant or a substance derived from the supernatant in any one of claims 205-244. . The dairy product or dairy product substitute in claim 245, wherein the dairy product or dairy product substitute is cheese. . A dairy product or dairy product substitute, comprising a recombinant casein protein and a coagulant. . The dairy product or dairy product substitute in claim 247, wherein the coagulant is in a detectable amount. . The dairy product or dairy product substitute in claim 247, wherein the coagulant is a magnesium salt. . The dairy product or dairy product substitute in claim 249, wherein the magnesium salt is at least one of magnesium chloride (MgCh) or magnesium sulfate (MgSC ). . The dairy product or dairy product substitute in claim 247, wherein the coagulant is a calcium salt. . The dairy product or dairy product substitute in claim 251, wherein the calcium salt is at least one of calcium chloride (CaCh) or calcium sulfate (CaSC ). . The dairy product or dairy product substitute in claim 247, wherein the coagulant is glucono-delta-1 actone (GDL). . The dairy product or dairy product substitute in any one of claims 247-253, wherein the coagulant is less than 0.1% (w/w) the cheese product. . The dairy product or dairy product substitute in any one of claims 247-253, wherein the coagulant is less than 0.05% (w/w) the cheese product. . The dairy product or dairy product substitute in any one of claims 247-253, wherein the coagulant is less than 0.01% (w/w) the cheese product. . The dairy product or dairy product substitute in any one of claims 247-253, wherein the coagulant is less than 0.005% (w/w) the cheese product. . The dairy product or dairy product substitute in any one of claims 247-253, wherein the coagulant is less than 0.001% (w/w) the cheese product. . The dairy product or dairy product substitute in any one of claims 247-253, wherein the coagulant is more than 0.0005% (w/w) the cheese product.

. The dairy product or dairy product substitute in any one of claims 247-253, wherein the coagulant is more than 0.0001% (w/w) the cheese product. . The dairy product or dairy product substitute in any one of claims 245-253, wherein the dairy product or dairy product substitute has a total fat content below 1%. . The dairy product or dairy product substitute in any one of claims 245-253, wherein the dairy product or dairy product substitute has a total fat content below 0.1%. . The dairy product or dairy product substitute in any one of claims 245-253, wherein the dairy product or dairy product substitute has a total fat content below 0.01%. . The dairy product or dairy product substitute in any one of claims 245-253, wherein the dairy product or dairy product substitute has no detectable amount of animal fat. . The dairy product or dairy product substitute in any one of claims 245-253, wherein the dairy product or dairy product substitute has no detectable amount of lactose. . The dairy product or dairy product substitute in any one of claims 245-253, wherein the dairy product or dairy product substitute has no detectable amount of animal hormone. . The dairy product or dairy product substitute in claim 266, wherein the animal hormone is at least one of estrogens, progesterone, corticoid, and androgen. . The dairy product or dairy product substitute in any one of claims 245-267, wherein the dairy product or dairy product substitute is at least one of milk, cheese, yogurt, or butter.. A composition comprising milk solids and soy proteins; wherein the soy proteins comprise glycinin and conglycinin, wherein the ratio of glycinin/conglycinin is elevated compared to a naturally occurring ratio, wherein the elevated glycinin/conglycinin ratio leads to better curd formation when the composition is subject to a curd forming condition. . A composition comprising milk solids and soy proteins; wherein the soy proteins comprise glycinin and conglycinin, wherein the ratio of glycinin/conglycinin is reduced compared to a naturally occurring ratio, wherein the reduced glycinin/conglycinin ratio leads to better curd formation when the composition is subject to a curd forming condition. . The composition in claim 269 or 270, wherein the weight ratio of the milk solids and the soy proteins is at least 60:40. . The composition in claim 269 or 270, wherein the weight ratio of the milk solids and the soy proteins is at least 65:35. . The composition in claim 269 or 270, wherein the weight ratio of the milk solids and the soy proteins is at least 70:30. . The composition in claim 269 or 270, wherein the weight ratio of the milk solids and the soy proteins is at least 75:25.

. The composition in claim 269 or 270, wherein the weight ratio of the milk solids and the soy proteins is at least 80:20. . The composition in any one of claims 269-275, wherein the naturally occurring ratio is between 0.5-1.3. . A composition, comprising: casein micelles; and a soy ingredient comprising at least one of 7s or 1 Is, wherein the weight or molar ratio of the 7s and the I ls (7s/l Is) is higher than a naturally occurring ratio. . The composition of claim 277, wherein the naturally occurring weight ratio of the 7s and the I ls (7s/l Is) is between 0.5-1.3. . The composition of claim 278, wherein the ratio of the 7s and the I ls (7s/l Is) is higher than 1.3. . The composition of claim 278, wherein the ratio of the 7s and the I ls (7s/l Is) is higher than 2. . The composition of claim 278, wherein the ratio of the 7s and the I ls (7s/l Is) is higher than 5. . The composition of claim 278, wherein the ratio of the 7s and the I ls (7s/l Is) is higher than 10. . The composition of claim 278, wherein the ratio of the 7s and the I ls (7s/l Is) is higher than 20. . The composition of claim 278, wherein the ratio of the 7s and the I ls (7s/l Is) is higher than 50. . The composition of claim 278, wherein the ratio of the 7s and the I ls (7s/l Is) is higher than 100. . The composition of claim 278, wherein the ratio of the 7s and the I ls (7s/l Is) is higher than 1000. . The composition of any one of claims 277-286, wherein the casein micelle comprises recombinant casein proteins. . A composition, comprising: casein micelles; and a soy ingredient comprising at least one of P-conglycinin (7 s) or glycinin (I ls), wherein the ratio of 11 s/7s is less than a naturally occurring ratio. . The composition of claim 288, wherein the casein micelle comprises recombinant casein proteins.