NZ727699B2 - Processes for producing industrial products from plant lipids - Google Patents

Abstract

The present invention relates to methods of producing industrial products from plant lipids, particularly from vegetative parts of plants. In particular, the present invention provides oil products such as biodiesel and synthetic diesel and processes for producing these, as well as plants having an increased level of one or more non- polar lipids such as triacylglycerols and an increased total non-polar lipid content. In one particular embodiment, the present invention relates to combinations of modifications in two or more of lipid handling enzymes, oil body proteins, decreased lipid catabolic enzymes and/or transcription factors regulating lipid biosynthesis to increase the level of one or more non-polar lipids and/or the total non-polar lipid content and/or mono-unsaturated fatty acid content in plants or any part thereof. In an embodiment, the present invention relates to a process for extracting lipids. In another embodiment, the lipid is converted to one or more hydrocarbon products in harvested plant vegetative parts to produce alkyl esters of the fatty acids which are suitable for use as a renewable biodiesel fuel.

Claims (31)

1. A transgenic plant, or part thereof, comprising a recombinant plant leaf or stem cell comprising a) a first exogenous polynucleotide which encodes a transcription factor polypeptide which is a Wrinkled 1 (WRI1) polypeptide, b) a second exogenous polynucleotide which encodes diacylglycerol acyltransferase (DGAT) or a phosphatidylcholine diacylglycerol acyltransferase (PDAT), c) a first genetic modification which down-regulates endogenous production and/or activity of a sugar-dependent lipase 1 (SDP 1), wherein each exogenous polynucleotide is operably linked to a promoter which is capable of directing expression of the polynucleotide in the cell, and wherein before the plant flowers the cell comprises a total non-polar lipid content of at least 8% (w/w dry weight).

2. The plant, or part thereof, of claim 1, wherein the cell further comprises one or both of d) a third exogenous polynucleotide which encodes a fatty acid thioesterase, and e) a fourth exogenous polynucleotide which encodes a second transcription factor selected from the group consisting of Leafy Cotyledon 1 (LEC1), LEC1-like, Leafy Cotyledon 2 (LEC2), BABY BOOM (BBM), FUS3, ABI3, ABI4, ABI5, Dof4 and Dof11.

3. The plant, or part thereof, of claim 1 or claim 2 which further comprises one or more or all of a) a fifth exogenous polynucleotide which encodes an oil body coating (OBC) polypeptide, b) a second genetic modification which down-regulates endogenous production and/or activity of a trigalactosyldiacylglycerol (TGD) polypeptide, and c) a third genetic modification which down-regulates endogenous production and/or activity of a polypeptide involved in diacylglycerol (DAG) production in the plastids of the cell when compared to a corresponding cell lacking the third genetic modification.

4. The plant, or part thereof, according to any one of claims 1 to 3, wherein the part is a vegetative part and one or more or all of the promoters are expressed at a higher level in the vegetative part relative to seed of the plant.

5. The plant, or part thereof, according to any one of claims 1 to 4, wherein one or more or all of the following features apply; i) the cell has an increased synthesis of total fatty acids relative to a corresponding cell lacking the first exogenous polynucleotide, or a decreased catabolism of total fatty acids relative to a corresponding cell lacking the first exogenous polynucleotide, or both, such that it has an increased level of total fatty acids relative to a corresponding cell lacking the first exogenous polynucleotide, ii) the cell has an altered ratio of C16:3 to C18:3 fatty acids in its total fatty acid content and/or its galactolipid content relative to a corresponding cell lacking the exogenous polynucleotide(s) and/or genetic modification(s), iii) the cell is in a vegetative part of a plant and comprises a total non-polar lipid content of between 8% and 75% (w/w dry weight), iv) the cell is in a vegetative part of a plant and comprises a TAG content of between 8% and 75% (w/w dry weight), v) the second transcription factor polypeptide is Leafy Cotyledon 2 (LEC2), vi) oleic acid comprises at least 20% (mol%) of the total fatty acid content in the cell, vii) one or more or all of the promoters are selected from a tissue-specific promoter, a developmentally regulated promoter, an inducible promoter, or a circadian-rhythm regulated promoter, viii) the cell comprises a total fatty acid content which comprises C12:0 or C14:0 medium chain fatty acids, at a level of at least 5% of the total fatty acid content, ix) the cell comprises a total fatty acid content whose oleic acid level is increased by at least 2% relative to a corresponding cell lacking the exogenous polynucleotide(s) and/or genetic modification(s), and/or whose ?-linolenic acid (ALA) level is decreased by at least 2% relative to a corresponding cell lacking the exogenous polynucleotide(s) and/or genetic modification(s), x) non-polar lipid in the cell comprises a modified level of total sterols, or free (non- esterified) sterols, steroyl esters, steroyl glycosides, relative to the non-polar lipid in a corresponding cell lacking the exogenous polynucleotide(s) and/or genetic modification(s), xi) the cell comprises an exogenous polynucleotide encoding a silencing suppressor, wherein the exogenous polynucleotide is operably linked to a promoter which is capable of directing expression of the polynucleotide in the cell, and xii) a total polyunsaturated fatty acid (PUFA) content which is decreased relative to the total PUFA content of a corresponding cell lacking the exogenous polynucleotide(s) and/or genetic modification(s).

6. The plant, or part thereof, according to any one of claims 1 to 5, wherein one or more or all of the following features apply; i) the second exogenous polynucleotide encodes a DGAT, ii) the oil body coating (OBC) polypeptide is oleosin or a lipid droplet associated protein (LDAP), iv) the fatty acid thioesterase is a C16 or C18 fatty acid thioesterase, v) the TGD polypeptide is a TGD5 polypeptide, and vi) the polypeptide involved in diacylglycerol (DAG) production in the plastid is a plastidial GPAT.

7. The plant, or part thereof, of claim 6, wherein the fatty acid thioesterase is a FATA polypeptide or a FATB polypeptide.

8. The plant, or part thereof, according to any one of claims to 1 to 7, wherein before the plant flowers the cell comprises a total non-polar lipid content of at least about 10%, at least about 11%, between 8% and 15%, or between 9% and 12% (w/w dry weight).

9. The plant, or part thereof, according to any one of claims 1 to 8, wherein the first genetic modification is a mutation of an endogenous SDP1 gene which partially or completely inactivates the gene, or the first genetic modification is an exogenous polynucleotide encoding an RNA molecule which inhibits expression of the endogenous SDP1 gene, wherein the exogenous polynucleotide is operably linked to a promoter which is capable of directing expression of the polynucleotide in the cell.

10. A population of at least about 1000 plants, each being a plant according to any one of claims 1 to 9, growing in a field, or a collection of at least about 1000 vegetative plant parts, each being a plant part according to any one of claims 1 to 9, wherein the vegetative plant parts have been harvested from plants growing in a field.

11. Seed of, or obtained from, a plant according to any one of claims 1 to 9.

12. A process for obtaining a transgenic plant according to any one of claims 1 to 9, the process comprising the steps of: i) introducing into a plant cell at least one exogenous polynucleotide and/or at least one genetic modification as defined in any one of claims 1 to 9, ii) producing a transgenic plant from the cell of step i), iii) expressing the exogenous polynucleotide(s) in the transgenic plant, iv) analysing the lipid content of at least a part of a leaf or stem of the transgenic plant, v) selecting a transgenic plant according to any one of claims 1 to 9.

13. A method of producing a plant which has integrated into its genome a set of exogenous polynucleotides and/or genetic modifications as defined in any one of claims 1 to 9, the method comprising the steps of i) screening one or more progeny plants obtained from a cross of two parental plants, wherein one plant comprises at least one of the exogenous polynucleotides and/or at least one genetic modification as defined in any one of claims 1 to 9, and the other plant comprises at least one of the exogenous polynucleotides and/or at least one genetic modifications as defined in any one of claims 1 to 9, and wherein between them the two parental plants comprise a set of exogenous polynucleotides and/or genetic modification as defined in any one of claims 1 to 9, for the presence or absence of the set of exogenous polynucleotides and/or genetic modifications as defined in any one of claims 1 to 9, and iii) selecting a progeny plant which comprises the set of exogenous polynucleotides and/or genetic modifications as defined in any one of claims 1 to 9, thereby producing the plant.

14. A process for producing an industrial product, the process comprising the steps of: i) obtaining a transgenic plant or part thereof of any one of claims 1 to 9, and ii) either a) converting at least some of the lipid in the plant or part thereof of step i) to the industrial product by applying heat, chemical, or enzymatic means, or any combination thereof, to the lipid in situ in the plant or part thereof, or b) physically processing the plant or part thereof of step i), and subsequently or simultaneously converting at least some of the lipid in the processed plant or part thereof to the industrial product by applying heat, chemical, or enzymatic means, or any combination thereof, to the lipid in the processed plant or part thereof, and iii) recovering the industrial product, thereby producing the industrial product.

15. The process of claim 14, wherein the plant part is a vegetative plant part.

16. The process of claim 14 or claim 15, further comprising steps of: (1) extracting at least some of the non-polar lipid content of the plant or part thereof as non-polar lipid, and (2) recovering the extracted non-polar lipid, wherein steps (1) and (2) are performed prior to the step of converting at least some of the lipid in the plant or part thereof to the industrial product.

17. A process for producing extracted lipid, the process comprising the steps of: i) obtaining a transgenic plant or part thereof of any one of claims 1 to 9, ii) extracting lipid from the plant or part thereof, and iii) recovering the extracted lipid, thereby producing the extracted lipid.

18. The process of claim 16 or claim 17 which comprises one or more of degumming, deodorising, decolourising, drying, fractionating the extracted lipid, removing wax esters from the extracted lipid, and analysing the fatty acid composition of the extracted lipid.

19. The process according to any one of claims 16 to 18, wherein the process further comprises converting the extracted lipid to an industrial product.

20. The process according to any one of claims 14 to 16 or 19, wherein the industrial product is a hydrocarbon product.

21. The process of claim 20, wherein the hydrocarbon product comprises a fatty acid ester, an alkane, a mixture of longer chain alkanes, an alkene, or a bioalcohol.

22. A process for producing seed, the process comprising: i) growing a plant according to any one of claims 1 to 9, and ii) harvesting seed from the plant.

23. A process for producing fuel, the process comprising reacting the lipid obtained by the process of claim 17 or claim 18 with an alcohol in the presence of a catalyst, to produce alkyl esters.

24. The process of claim 23 which further comprises blending the alkyl esters with petroleum-based fuel.

25. A process for producing a synthetic diesel fuel, the process comprising: i) converting the lipid in a transgenic plant or part thereof of any one of claims 1 to 9 to a bio-oil by a process comprising pyrolysis or hydrothermal processing or to a syngas by gasification, and ii) converting the bio-oil to synthetic diesel fuel by a process comprising fractionation, or converting the syngas to a biofuel using a metal catalyst or a microbial catalyst.

26. A process for producing a biofuel, the process comprising converting the transgenic plant or part thereof of any one of claims 1 to 9 to bio-oil by pyrolysis, a bioalcohol by fermentation, or a biogas by gasification or anaerobic digestion.

27. The process of claim 25 or claim 26, wherein the part is a vegetative plant part.

28. A process for producing a feedstuff, the process comprising admixing a transgenic plant or part thereof of any one of claims 1 to 9 with at least one other food ingredient.

29. A feedstuff comprising a transgenic plant or part thereof of any one of claims 1 to 9.

30. A process for feeding an animal, the process comprising providing to the animal a transgenic plant or part thereof of any one of claims 1 to 9.

31. A process for producing an oil product, the process comprising the steps of (i) treating, in a reactor, a composition comprising (a) plant parts according to any one of claims 1 to 10 which are vegetative plant parts whose dry weight is at least 2g and which have a total non-polar lipid content of at least 5% by weight on a dry weight basis, and (b) a solvent which comprises water, an alcohol, or both, wherein the treatment comprises heating the composition at a temperature between about 50 C and about 450 C and at a pressure between 5 and 350 bar for between 1 and