[go: up one dir, main page]

WO2024161358A1 - Résistance au virus de la jaunisse des betteraves - Google Patents

Résistance au virus de la jaunisse des betteraves Download PDF

Info

Publication number
WO2024161358A1
WO2024161358A1 PCT/IB2024/050952 IB2024050952W WO2024161358A1 WO 2024161358 A1 WO2024161358 A1 WO 2024161358A1 IB 2024050952 W IB2024050952 W IB 2024050952W WO 2024161358 A1 WO2024161358 A1 WO 2024161358A1
Authority
WO
WIPO (PCT)
Prior art keywords
plant
seq
chromosome
dna
refbeet
Prior art date
Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
Ceased
Application number
PCT/IB2024/050952
Other languages
English (en)
Inventor
Charlotte Britt-Louise LENNEFORS
Hans Mårten Joakim HERRSTRÖM
Current Assignee (The listed assignees may be inaccurate. Google has not performed a legal analysis and makes no representation or warranty as to the accuracy of the list.)
DLF Seeds AS
Original Assignee
DLF Seeds AS
Priority date (The priority date is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the date listed.)
Filing date
Publication date
Application filed by DLF Seeds AS filed Critical DLF Seeds AS
Priority to EP24703866.4A priority Critical patent/EP4658061A1/fr
Publication of WO2024161358A1 publication Critical patent/WO2024161358A1/fr
Anticipated expiration legal-status Critical
Ceased legal-status Critical Current

Links

Classifications

    • A—HUMAN NECESSITIES
    • A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
    • A01H—NEW PLANTS OR NON-TRANSGENIC PROCESSES FOR OBTAINING THEM; PLANT REPRODUCTION BY TISSUE CULTURE TECHNIQUES
    • A01H6/00—Angiosperms, i.e. flowering plants, characterised by their botanic taxonomy
    • A01H6/02—Amaranthaceae or Chenopodiaceae, e.g. beet or spinach
    • A—HUMAN NECESSITIES
    • A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
    • A01H—NEW PLANTS OR NON-TRANSGENIC PROCESSES FOR OBTAINING THEM; PLANT REPRODUCTION BY TISSUE CULTURE TECHNIQUES
    • A01H1/00—Processes for modifying genotypes ; Plants characterised by associated natural traits
    • A01H1/04—Processes of selection involving genotypic or phenotypic markers; Methods of using phenotypic markers for selection
    • A01H1/045—Processes of selection involving genotypic or phenotypic markers; Methods of using phenotypic markers for selection using molecular markers

Definitions

  • the present disclosure relates to plants, plant parts, and plant cells conferring traits of resistance to virus yellows and methods of producing the same. Further, the disclosure relates to nucleic acid molecules for identifying resistant traits, and nucleic acid molecules that impart resistance to pathogens in plants, plant parts, and plant cells.
  • sequence listing xml associated with this application is provided electronically in xml file format and is hereby incorporated by reference into the specification.
  • the contents of the electronic sequence listing (DLFS_001_01WO_SeqList_ST26.xml; Size: 77,623 bytes; and Date of Creation: January 26, 2024) are herein incorporated by reference in its entirety.
  • the beet yellowing disease appears in circles in the fields, in the form of lightening and then yellowing of the lamina between the veins of leaves.
  • the leaves thicken and become brittle .
  • These symptoms first form limited areas of infection in the fields and then rapidly spread throughout the entire field. Yellowing viruses can cause yield losses of 50 % when it infects the crop in early June. Infection reduces the photosynthetic area of leaves reducing yield and sugar content, which significantly impacts the world’s sugar production and related food chains in human beings and animals.
  • the disease is widespread across all sugar beet regions of Europe. If it strikes early in the season, it can be the cause of significant yield loss, reduced sugar content and lower industrial quality.
  • the present disclosure relates to a. Beta vulgaris subsp. vulgaris plant, plant part, or plant cell resistant to Beet Mild Yellowing Virus (BMYV), wherein said plant, plant part, or plant cell includes at least one Quantitative Trait Loci (QTL) associated with resistance to BMYV, wherein said QTL is selected from the group consisting of QTL1, QTL2, and QTL3, and wherein QTL1, QTL2, and QTL 3, are obtainable from Beta vulgaris subsp. vulgaris line 21000003-71, representative seed of which has been deposited under NCIMB number 44107.
  • QTL Quantitative Trait Loci
  • the present disclosure relates to a Beta vulgaris subsp. vulgaris plant, plant part, or plant cell, wherein QTL1 is genetically linked to at least one single nucleotide polymorphism (SNP) selected from the group consisting of PBV504245, PBV125552, PBV291217, PBV369685, and PBV261963, wherein: PBV504245 is a C to G substitution corresponding to position 1050427 of chromosome 1 of the Refbeet 1.5 reference genome, PBV125552 is a G to A substitution corresponding to position 3069972 of chromosome 1 of the Refbeet 1.5 reference genome, PBV291217 is a G to A substitution corresponding to position 4315236 of chromosome 1 of the Refbeet 1.5 reference genome, PBV369685 is an A to G substitution corresponding to position 4330737 of chromosome 1 of the Refbeet 1.5 reference genome, and PBV261963 is an Ato
  • SNP single nucle
  • QTL1 is genetically linked to at least one single nucleotide polymorphism (SNP) selected from the group consisting of PBV757437, PBV638722, and EPBV6296, wherein: PBV757437 is a G to A substitution corresponding to position 17043 of chromosome 1 of the Refbeet 1.5 reference genome, PBV638722 is an A to G substitution corresponding to position 2645728 of chromosome 1 of the Refbeet 1.5 reference genome, and EPBV6296 is a T to C substitution corresponding to position 1117040 of chromosome 1 of the Refbeet 1.5 reference genome.
  • SNP single nucleotide polymorphism
  • the present disclosure relates to a. Beta vulgaris subsp. vulgaris plant, plant part, or plant cell, wherein QTL2 is genetically linked to at least one single nucleotide polymorphism (SNP) selected from the group consisting of PBV577030, PBV562771, PBV573138, PBV273480, and PBV821051, wherein: PBV577030 is an A to G substitution corresponding to position 5742753 of chromosome 4 of the Refbeet 1.5 reference genome, PBV562771 is a G to A substitution corresponding to position 9767085 of chromosome 4 of the Refbeet 1.5 reference genome, PBV573138 is an A to G substitution corresponding to position 10271308 of chromosome 4 of the Refbeet 1.5 reference genome, PBV273480 is a G to A substitution corresponding to position 46718199 of chromosome 4 of the Refbeet 1.5 reference genome, and PBV821051 is
  • the present disclosure relates to a Beta vulgaris subsp. vulgaris plant, plant part, or plant cell, wherein QTL3 is genetically linked to at least one single nucleotide polymorphism (SNP) selected from the group consisting of PBV148969, PBV331644, PBV882826, PBV374371, and PBV757436, wherein: PBV148969 is an A to G substitution corresponding to position 436005 of chromosome 8 of the Refbeet 1.5 reference genome, PBV331644 is a G to A substitution corresponding to position 1366650 of chromosome 8 of the Refbeet 1.5 reference genome, PBV882826 is an A to G substitution corresponding to position 7979633 of chromosome 8 of the Refbeet 1.5 reference genome, PBV374371 is a G to A substitution corresponding to position 8822544 of chromosome 8 of the Refbeet 1.5 reference genome, and PBV757436 is an A to
  • QTL3 is genetically linked to at least one single nucleotide polymorphism (SNP) selected from the group consisting of PBV738292, PBV738293, and EPBV7349, wherein: PBV738292 is a Gto an A substitution corresponding to position 35831 of chromosome 8 of the Refbeet 1.5 reference genome, PBV738293 is an A to G substitution corresponding to position 1366650 of chromosome 8 of the Refbeet 1.5 reference genome, and EPBV7349 is an A to C substitution corresponding to position 275340 of chromosome 8 of the Refbeet 1.5 reference genome.
  • SNP single nucleotide polymorphism
  • the present disclosure relates to a. Beta vulgaris subsp. vulgaris plant, plant part, or plant cell, further including a fourth QTL (QTL4) associated with resistance to BMYV, wherein QTL4 is genetically linked to at least one marker loci selected from the group consisting of GRKPBV98281, GRKPBV627372, GRKPBV672273, GRKPBV627178, GRKPBV 116271, and EPBV8821, wherein: GRKPBV98281 is a G to A substitution corresponding to position 8040503 of chromosome 2 of the Refbeet 1.5 reference genome, GRKPBV627372 is a C to an A substitution corresponding to position 9433027 of chromosome 2 of the Refbeet 1.5 reference genome, GRKPBV672273 is a G to an A substitution corresponding to position 12212298 of chromosome 2 of the Refbeet 1.5 reference genome, GRK
  • the present disclosure relates to a plant, plant part, or plant cell having resistance to Beet Mild Yellows Virus, wherein the resistance is associated with a reduced load of the Beet Mild Yellows Virus by at least 25% in comparison to a variety lacking QTL1, QTL2, QTL3, and/or QTL4. In some aspects, the resistance is associated with a reduced load of the Beet Mild Yellows Virus by at least 50% in comparison to a variety lacking QTL1, QTL2, QTL3, and/or QTL4. In some aspects, the resistance is associated with a reduced load of the Beet Mild Yellows Virus by at least 75% in comparison to a variety lacking QTL1, QTL2, QTL3, and/or QTL4.
  • the reduced load of the Beet Mild Yellows Virus is determined by an antibody assay.
  • the antibody assay is an enzyme-linked immunosorbent assay.
  • the reduced load of the Beet Mild Yellows Virus is determined by an antibody assay in comparison to a plant, plant part, or plant cell which is genetically essentially identical except for the presence of the resistance to BMYV.
  • the plant, plant part, or plant cell is also resistant against Beet yellows virus (BYV) or Beet chlorosis virus (BChV).
  • the present disclosure relates to a plant, plant part, or plant cell, wherein said plant, plant part, or plant cell is heterozygous for at least one of QTL1, QTL2, QTL3, and QTL4.
  • the plant, plant part, or plant cell is heterozygous for QTL1 and QTL2.
  • the plant, plant part, or plant cell is heterozygous for QTL1 and QTL3.
  • the plant, plant part, or plant cell is heterozygous for QTL1 and QTL4.
  • the plant, plant part, or plant cell is heterozygous for QTL2 and QTL3.
  • the plant, plant part, or plant cell is heterozygous for QTL2 and QTL4.
  • the plant, plant part, or plant cell is heterozygous for QTL3 and QTL4.
  • the plant, plant part, or plant cell is heterozygous for QTL1, QTL2, QTL3, and QTL4.
  • the present disclosure relates to a plant, plant part, or plant cell, wherein said plant, plant part, or plant cell is homozygous for at least one of QTL1, QTL2, QTL3, and QTL4.
  • the plant, plant part, or plant cell is homozygous for QTL1 and QTL2.
  • the plant, plant part, or plant cell is homozygous for QTL1 and QTL3.
  • the plant, plant part, or plant cell is homozygous for QTL3 and QTL4.
  • the plant, plant part, or plant cell is homozygous for QTL2 and QTL3.
  • the plant, plant part, or plant cell is homozygous for QTL2 and QTL4.
  • the plant, plant part, or plant cell is homozygous for QTL3 and QTL4.
  • the plant, plant part, or plant cell is homozygous for QTL1, QTL2, QTL3, and QTL4.
  • the present disclosure relates to a plant, plant part, or plant cell, wherein said plant, plant part, or plant cell is heterozygous for QTL1, and homozygous for at least one of QTL2, QTL3, and QTL4.
  • the plant, plant part, or plant cell is heterozygous for QTL2, and homozygous for at least one of QTL1, QTL3, and QTL4.
  • the plant, plant part, or plant cell is heterozygous for QTL3, and homozygous for at least one of QTL1, QTL2, and QTL4.
  • the plant, plant part, or plant cell is heterozygous for QTL4, and homozygous for at least one of QTL1, QTL2, and QTL3.
  • the plant, plant part, or plant cell is homozygous for QTL1, and heterozygous for at least one of QTL2, QTL3, and QTL4. In some aspects, the plant, plant part, or plant cell is homozygous for QTL2, and heterozygous for at least one of QTL1, QTL3, and QTL4. In some aspects, the plant, plant part, or plant cell is homozygous for QTL3, and heterozygous for at least one of QTL1, QTL2, and QTL4. In some aspects, the plant, plant part, or plant cell is homozygous for QTL4, and heterozygous for at least one of QTL1, QTL2, and QTL3.
  • the present disclosure relates to a plant, plant part, or plant cell as described herein, wherein said plant is a fodder beet or sugar beet.
  • the present disclosure relates to a plant, plant part, or plant cell plant as described herein, wherein the plant part, or plant cell includes a desirable trait introduced by a method selected from the group consisting of genetic transformation, genome editing, targeted mutagenesis, and induced random mutagenesis.
  • the desirable trait is selected from the group consisting of (i) the H7-1 event conferring glyphosate tolerance, (ii) a mutated ALS gene conferring tolerance against ALS herbicides, (iii) the GTSB77 event conferring glyphosate tolerance, and (iv) the T 120-7 event conferring glufosinate tolerance.
  • the ALS gene encodes an ALS polypeptide containing an amino acid different from tryptophan at a position 569 of the ALS polypeptide.
  • the plant, plant part, or plant cell further includes a Beet Mild Yellowing Virus tolerance trait obtainable from the group of sugar beet varieties including Maruscha KWS and Novalina KWS.
  • the present disclosure relates to a plant part as described herein, wherein the plant part is a seed.
  • the seed is technically treated, whereby the technical treatment is selected from the group consisting of polishing, pelleting, incrustation, and coloring.
  • the technical treatment does not include insecticide.
  • the plant part is a commodity plant product.
  • the plant product is a beet.
  • the present disclosure relates to a marker for selection of Beet Mild Yellowing Virus resistant plants selected from the group consisting of SEQ ID NO: 1 to SEQ ID NO: 45. In some aspects, the present disclosure relates to a marker for selection of Beet Mild Yellowing Virus resistant plants selected from the group consisting of SEQ ID NO: 52 to SEQ ID NO: 87.
  • the marker utilizes a sequence selected from the group consisting of SEQ ID NO: 2, SEQ ID NO: 5, SEQ ID NO: 8, SEQ ID NO: 11, SEQ ID NO: 14, SEQ ID NO: 17, SEQ ID NO: 20, SEQ ID NO: 23, SEQ ID NO: 26, SEQ ID NO: 29, SEQ ID NO: 32, SEQ ID NO: 35, SEQ ID NO: 38, SEQ ID NO: 41, and SEQ ID NO: 44.
  • the marker utilizes a sequence selected from the group consisting of SEQ ID NO: 52, SEQ ID NO: 56, SEQ ID NO: 58, SEQ ID NO: 62, SEQ ID NO: 65, SEQ ID NO: 67, SEQ ID NO: 70, SEQ ID NO: 73, SEQ ID NO: 76, SEQ ID NO: 79, SEQ ID NO: 83, and SEQ ID NO: 85.
  • the present disclosure relates to a method of using any one of the molecular markers described herein for selection of a plant resistant against a virus selected from the group consisting of Beet Mild Yellowing Virus (BMYV), Beet yellows virus (BYV), and Beet chlorosis virus (BChV).
  • BMYV Beet Mild Yellowing Virus
  • BYV Beet yellows virus
  • BChV Beet chlorosis virus
  • the molecular markers identify at least one of the Single Nucleotide Polymorphisms present in PBV504245, PBV125552, PBV291217, PBV369685, PBV261963, PBV577030, PBV562771, PBV573138, PBV273480, PBV821051, PBV148969, PBV331644, PBV882826, PBV374371, and PBV757436. In some aspects, the molecular markers identify at least one of the Single Nucleotide Polymorphisms present in PBV757437, PBV638722, EPBV6296, PBV738292, PBV738293, EPBV7349,
  • the techniques described herein relate to a method of identifying a plant, plant part, or plant cell including a Quantitative Trait Locus (QTL) associated with resistance to Beet Mild Yellow Virus (BMYV) on chromosome 1 (QTL1), said method including the steps of: screening for the presence of QTL1, wherein QTL1 is genetically linked to at least one single nucleotide polymorphism (SNP) selected from the group consisting of PBV504245, PBV125552, PBV291217, PBV369685, and PBV261963, wherein: PBV504245 is a C to G substitution corresponding to position 1050427 of chromosome 1 of the Refbeet 1.5 reference genome, PBV125552 is a G to A substitution corresponding to position 3069972 of chromosome 1 of the Refbeet 1.5 reference genome, PBV291217 is a G to A substitution corresponding to position 4315236 of chromosome 1 of the
  • QTL1 is genetic
  • the screening for the presence of QTL1 further includes identifying a plant, plant part, or plant cell having at least one single nucleotide polymorphism (SNP) selected from the group consisting of PBV757437, PBV638722, and EPBV6296, wherein: PBV757437 is a G to A substitution corresponding to position 17043 of chromosome 1 of the Refbeet 1.5 reference genome, PBV638722 is an A to G substitution corresponding to position 2645728 of chromosome 1 of the Refbeet 1.5 reference genome, and EPBV6296 is a T to C substitution corresponding to position 1117040 of chromosome 1 of the Refbeet 1.5 reference genome.
  • SNP single nucleotide polymorphism
  • the screening for the presence of QTL1 includes at least one of: PCR amplification of DNA with SEQ ID NO: 2 and SEQ ID NO: 3, PCR amplification of DNA with SEQ ID NO: 5 and SEQ ID NO: 6, PCR amplification of DNA with SEQ ID NO: 8 and SEQ ID NO: 9, PCR amplification of DNA with SEQ ID NO: 11 and SEQ ID NO: 12, and PCR amplification of DNA with SEQ ID NO: 14 and SEQ ID NO: 15.
  • the screening for the presence of QTL1 includes at least one of: PCR amplification of DNA with SEQ ID NO: 52 and SEQ ID NO: 54, PCR amplification of DNA with SEQ ID NO: 56 and SEQ ID NO: 57, and PCR amplification of DNA with SEQ ID NO: 58 and SEQ ID NO: 60.
  • the techniques described herein relate to a method of identifying a plant, plant part, or plant cell including a Quantitative Trait Locus (QTL) associated with resistance to Beet Mild Yellow Virus (BMYV) on chromosome 4 (QTL2), said method including the steps of: screening for the presence of QTL2, wherein QTL2 is genetically linked to at least one single nucleotide polymorphism (SNP) selected from the group consisting of PBV577030, PBV562771, PBV573138, PBV273480, and PBV82I05I, wherein: PBV577030 is an A to G substitution corresponding to position 5742753 of chromosome 4 of the Refbeet 1.5 reference genome, PBV562771 is a G to A substitution corresponding to position 9767085 of chromosome 4 of the Refbeet 1.5 reference genome, PBV573138 is an A to G substitution corresponding to position 10271308 of chromosome 4
  • QTL2 is genetic
  • the screening for the presence of QTL2 includes at least one of: PCR amplification of DNA with SEQ ID NO: 17 and SEQ ID NO: 18, PCR amplification of DNA with SEQ ID NO: 20 and SEQ ID NO: 21, PCR amplification of DNA with SEQ ID NO: 23 and SEQ ID NO: 24, PCR amplification of DNA with SEQ ID NO: 26 and SEQ ID NO: 27, and PCR amplification of DNA with SEQ ID NO: 29 and SEQ ID NO: 30.
  • the techniques described herein relate to a method of identifying a plant, plant part, or plant cell including a Quantitative Trait Locus (QTL) associated with resistance to Beet Mild Yellow Virus (BMYV) on chromosome 8 (QTL3), said method including the steps of: screening for the presence of QTL3, wherein QTL3 is genetically linked to at least one single nucleotide polymorphism (SNP) selected from the group consisting of PBV148969, PBV331644, PBV882826, PBV374371, and PBV757436, wherein: PBV148969 is an A to G substitution corresponding to position 436005 of chromosome 8 of the Refbeet 1.5 reference genome, PBV331644 is a G to A substitution corresponding to position 1366650 of chromosome 8 of the Refbeet 1.5 reference genome, PBV882826 is an A to G substitution corresponding to position 7979633 of chromosome 8 of the Ref
  • QTL3 is genetic
  • the screening for the presence of QTL3 further includes identifying a plant, plant part, or plant cell having at least one single nucleotide polymorphism (SNP) selected from the group consisting of PBV738292, PBV738293, and EPBV7349, wherein: PBV738292 is a G to an A substitution corresponding to position 35831 of chromosome 8 of the Refbeet 1.5 reference genome, PBV738293 is an A to G substitution corresponding to position 1366650 of chromosome 8 of the Refbeet 1.5 reference genome, and EPBV7349 is an A to C substitution corresponding to position 275340 of chromosome 8 of the Refbeet 1.5 reference genome.
  • SNP single nucleotide polymorphism
  • the screening for the presence of QTL3 includes at least one of: PCR amplification of DNA with SEQ ID NO: 32 and SEQ ID NO: 33, PCR amplification of DNA with SEQ ID NO: 35 and SEQ ID NO: 36, PCR amplification of DNA with SEQ ID NO: 38 and SEQ ID NO: 39, PCR amplification of DNA with SEQ ID NO: 41 and SEQ ID NO: 42, and PCR amplification of DNA with SEQ ID NO: 44 and SEQ ID NO: 45.
  • the screening for the presence of QTL3 includes at least one of: PCR amplification of DNA with SEQ ID NO: 62 and SEQ ID NO: 63 PCR amplification of DNA with SEQ ID NO: 65 and SEQ ID NO: 66, and PCR amplification of DNA with SEQ ID NO: 67 and SEQ ID NO: 69.
  • the techniques described herein relate to a method of identifying a plant, plant part, or plant cell including a Quantitative Trait Locus (QTL) associated with resistance to Beet Mild Yellow Virus (BMYV) on chromosome 2 (QTL4), said method including the steps of: screening for the presence of QTL4, wherein QTL4 is genetically linked to at least one single nucleotide polymorphism (SNP) selected from the group consisting of GRKPBV98281, GRKPBV627372, GRKPBV672273, GRKPBV627178, GRKPBV116271, and EPBV8821, wherein: GRKPBV98281 is a G to A substitution corresponding to position 8040503 of chromosome 2 of the Refbeet 1.5 reference genome, GRKPBV627372 is a C to an A substitution corresponding to position 9433027 of chromosome 2 of the Refbeet 1.5 reference genome,
  • the screening for the presence of QTL4 includes at least one of: PCR amplification of DNA with SEQ ID NO: 70 and SEQ ID NO: 72, PCR amplification of DNA with SEQ ID NO: 73 and SEQ ID NO: 75, PCR amplification of DNA with SEQ ID NO: 76 and SEQ ID NO: 78, PCR amplification of DNA with SEQ ID NO: 79 and SEQ ID NO: 81, PCR amplification of DNA with SEQ ID NO: 83 and SEQ ID NO: 84, and PCR amplification of DNA with SEQ ID NO: 85 and SEQ ID NO: 87.
  • the techniques described herein relate to a method of producing a Beta vulgaris subsp. vulgaris plant, plant part, or plant cell having resistance to Beet Mild Yellowing Virus (BMYV), said method including the steps of: (a) providing a plant having resistance to Beet Mild Yellowing Virus (BMYV); (b) crossing the plant of (a) with another plant and harvesting the seed resulting from said cross; (c) growing the seed from step (b) to produce a progeny plant, and submitting the progeny plant to the method of identifying a plant including at least one of a Quantitative Trait Locus (QTL) associated with resistance to Beet Mild Yellow Virus (BMYV); and (d) selecting a progeny plant which includes at least one of QTL1, QTL2, and QTL3.
  • QTL Quantitative Trait Locus
  • the selecting further includes selecting a progeny plant having QTL4.
  • the selecting a progeny includes at least one of: PCR amplification of DNA with SEQ ID NO: 2 and SEQ ID NO: 3, PCR amplification of DNA with SEQ ID NO: 5 and SEQ ID NO: 6, PCR amplification of DNA with SEQ ID NO: 8 and SEQ ID NO: 9, PCR amplification of DNA with SEQ ID NO: 11 and SEQ ID NO: 12, and PCR amplification of DNA with SEQ ID NO: 14 and SEQ ID NO: 15.
  • the selecting a progeny includes at least one of: PCR amplification of DNA with SEQ ID NO: 52 and SEQ ID NO: 54, PCR amplification of DNA with SEQ ID NO: 56 and SEQ ID NO: 57, and PCR amplification of DNA with SEQ ID NO: 58 and SEQ ID NO: 60.
  • the selecting a progeny includes at least one of: PCR amplification of DNA with SEQ ID NO: 17 and SEQ ID NO: 18, PCR amplification of DNA with SEQ ID NO: 20 and SEQ ID NO: 21, PCR amplification of DNA with SEQ ID NO: 23 and SEQ ID NO: 24, PCR amplification of DNA with SEQ ID NO: 26 and SEQ ID NO: 27, and PCR amplification of DNA with SEQ ID NO: 29 and SEQ ID NO: 30.
  • the selecting a progeny includes at least one of: PCR amplification of DNA with SEQ ID NO: 32 and SEQ ID NO: 33, PCR amplification of DNA with SEQ ID NO: 35 and SEQ ID NO: 36, PCR amplification of DNA with SEQ ID NO: 38 and SEQ ID NO: 39, PCR amplification of DNA with SEQ ID NO: 41 and SEQ ID NO: 42, and PCR amplification of DNA with SEQ ID NO: 44 and SEQ ID NO: 45.
  • the selecting a progeny includes at least one of: PCR amplification of DNA with SEQ ID NO: 62 and SEQ ID NO: 63 PCR amplification of DNA with SEQ ID NO: 65 and SEQ ID NO: 66, and PCR amplification of DNA with SEQ ID NO: 67 and SEQ ID NO: 69.
  • the selecting a progeny includes at least one of: PCR amplification of DNA with SEQ ID NO: 70 and SEQ ID NO: 72, PCR amplification of DNA with SEQ ID NO: 73 and SEQ ID NO: 75, PCR amplification of DNA with SEQ ID NO: 76 and SEQ ID NO: 78, PCR amplification of DNA with SEQ ID NO: 79 and SEQ ID NO: 81, PCR amplification of DNA with SEQ ID NO: 83 and SEQ ID NO: 84, and PCR amplification of DNA with SEQ ID NO: 85 and SEQ ID NO: 87.
  • the method further includes (e) backcrossing said selected progeny plant to a parental line to produce a backcross progeny having two or more QTLs associated with resistance to Beet Mild Yellowing Virus. In some aspects, the method further includes (e) backcrossing said selected progeny plant to a parental line to produce backcross progeny having a desirable trait and resistance to Beet Mild Yellowing Virus. In some aspects, the method further includes introducing or introgressing a desirable trait introduced by a method selected from the group consisting of genetic transformation, genome editing, targeted mutagenesis, and induced random mutagenesis.
  • the present disclosure provides methods for reducing yield loss as a consequence of Beet Mild Yellowing Virus (BMYV) infection, the method comprising the steps of: (a) providing a seed for a plant having resistance to Beet Mild Yellowing Virus (BMYV), a plant identified by the method taught herein, or a plant produced by the method taught herein; and (b) planting said seed in areas which are prone to Beet Mild Yellowing Virus infection.
  • the seed is planted in areas where the use of insecticides to control the vector for Beet Mild Yellowing Virus transmission is restricted, prohibited, or not desired.
  • the present disclosure provides methods for producing sugar, the method comprising the steps of: (a) growing the plant having resistance to Beet Mild Yellowing Virus (BMYV), a plant identified by the method taught herein, or a plant produced by the method taught herein, (b) harvesting the root/beet of said plant, and (c) extracting the sugar from said root/beet of (b).
  • the growing is conducted without the use of insecticides and the sugar extracted from said root/beet is qualified for the organic market.
  • FIGs 1A- IF illustrate one embodiment of the present disclosure.
  • FIGs. 1A to IF illustrate the development of symptoms scored every 3 rd week during the growing season using a scale from 1-9, with 1 being the worst and 9 being the best. Examples of the scoring criteria are presented in FIG. 1A (score 3), FIG. IB (score 4), FIG. 1C (score 5), FIG. ID (score 6), FIG. IE (score 7), and FIG. IF (score 8).
  • the disclosure provides plants, plant parts, and plant cells that are resistant to yellowing viruses, and methods for producing plants exhibiting resistance to yellowing diseases caused by the pathogens such as Beet mild yellowing virus (BMYV), Beet yellows virus (BYV), Beet chlorosis virus (BChV), Beet mosaic virus (BtMV), and/or the Beet Western Yellows Virus (BWYV), thereby preventing yield loss and drag, and maintaining or even increasing yield for sugar production.
  • Methods of breeding and selecting multiple virus resistant sugar beet lines are further provided, as well as plants, seeds, and commodity plant products of the same.
  • molecular markers that are linked to quantitative trait loci contributing to such yellowing virus resistance.
  • the inventors have identified QTL and markers associated with said QTL and have developed plants and methods of producing plants and commodity plant products having resistance to yellowing viruses, such as BMYV, BYV, BChV, BtMV, and BWYW.
  • yellowing viruses such as BMYV, BYV, BChV, BtMV, and BWYW.
  • Signs of viral infection include but are not limited to, (i) yellow and brown in the area of older leaves between the veins where small reddish brown spots often appear, (ii) thick, leathery, and brittle leaves, (iii) yellow etching of the heart leaves, (iv) sugar yield loss in root, and the like.
  • the disclosure provides methods and compositions that permit combination of multivirus resistance with the ability to produce a commercially acceptable sugar beet from a single line.
  • the disclosure represents a significant advance in that it provides plants and methods of introgressing resistant traits into commercially acceptable genetic backgrounds that may be susceptible to the viruses or any susceptible plant line that is desired to acquire resistance to yellowing disease(s).
  • a QTL conferring BMYV resistance is identified and defined by at least marker taught herein.
  • QTL Quantitative Trait Loci
  • QTL1 located on chromosome 1
  • QTL2 located on chromosome 4
  • QTL3 located on chromosome 8
  • QTL4 located on chromosome 2
  • the first QTL (QTL1) identified herein that confers resistance to BMYV is located on chromosome 1 in the interval bounded by markers PBV504245, PBV125552, PBV291217, PBV369685, PBV261963, PBV757437, PBV638722, and EPBV6296.
  • the second QTL (QTL2) identified herein that confers resistance to BMYV is located on chromosome 4 in the interval bounded by markers PBV577030, PBV562771, PBV573138, PBV273480, and PBV821051.
  • the third QTL (QTL3) identified herein that confers resistance to BMYV is located on chromosome 8 in the interval bounded by markers PBV148969, PBV331644, PBV882826, PBV374371, PBV757436, PBV738292, PBV738293, and EPBV7349.
  • the fourth QTL (QTL4) identified herein that confers resistance to BMYV is located on chromosome 2 in the interval bounded by markers GRKPBV98281, GRKPBV627372, GRKPBV672273, GRKPBV627178, GRKPBV116271, and EPBV8821.
  • corresponding markers provided herein and/or other markers that may be linked thereto, one of skill in the art may use genetic markers to introgress and transfer virus resistance traits in commercially relevant varieties and beet lines.
  • identified QTL may be introgressed into any different Beta vulgaris subsp. vulgaris (beet) genetic background.
  • a beet plant of any genotype may be produced that further comprises the desired viral resistance, including BMYV, BYV and BChV.
  • such plants may be prepared to comprise other desired traits, for example elite agronomic and root quality traits as desired.
  • nucleic acid can mean that at least one nucleic acid can be utilized.
  • Beta vulgaris subsp. vulgaris plant includes species used for feed and industrial purpose such as sugar beet or a fodder beet, and species used for food purpose such as red beet and mangold.
  • plant part refers to cells, tissues or organs, seed pods, seeds, severed parts such as roots, leaves, flowers, pollen, root tips, anthers, pistils, embryos, ovules, cotyledons, hypocotyls, petiole, meristematic cells, shoots, gametophytes, sporophytes, and the like etc.
  • Progeny or descendants of the plants described herein which retain the distinguishing characteristics of the parents such as seed obtained by selfing or crossing, e.g. hybrid seed (obtained by crossing two inbred parental lines), hybrid plants and plant parts derived therefrom are encompassed herein, unless otherwise indicated.
  • progeny or “descendent” is intended to mean the offspring or the first and all further descendants from a cross with a plant of the disclosure that shows the resistance trait(s) and/or carries the genetic determinant(s) underlying the trait(s).
  • Progeny of the disclosure comprises descendants of any cross with a plant of the disclosure that carries the genetic determinant(s) causing the resistance trait(s).
  • Fl population or “Fl generation” or “Fl” as used herein refers to the first filial generation produced by a cross.
  • F2 population or “F2 generation” or “F2” as used herein refers to offspring produced by self-pollination of individuals of an Fl generation.
  • variable refers to a plant genotype that is distinct, stable and uniform in its characteristics when propagated.
  • Yellow Disease refers to the disease caused by the pathogens Beet Mild Yellowing Virus (BMYV), Beet Yellows Virus (BYV) and/or Beet Chlorosis Virus (BChV), a common disease affecting plants of the Chenopodieae family that presents with symptoms including yellowing and loss of photosynthetic ability, which can eventually lead to severe sugar yield losses due to reduced root size and sugar content.
  • BMYV Mild Yellowing Virus
  • BYV Beet Yellows Virus
  • BChV Beet Chlorosis Virus
  • BMYV resistance refers to resistance to one or more Beet Mild Yellowing Virus isolates Said resistance refers to a reduction of viral load and/or a plants ability to prevent and/or limit viral replication, resulting in a reducing in damage caused by BMYV infection compared to control plants not having resistance. Damage can be assessed as, for example, visual yellowing of the leaves, root size, sugar content and virus content. In particular, a reduction in damage is manifested in a reduced yield loss and/or loss off sugar productivity when plants are grown under disease pressure in the field, compared to control plants. Said resistance may also refer to plants that are completely resistant, i.e., plants on which no disease symptoms are found.
  • BMYV resistance can be assessed using a scale from 1 to 9, where 1 would indicate a completely dead plant, 2-3 severely yellowing plants, 4-6 moderately yellowing plants, 7-8 mildly yellowing plants, and 9 which would indicate a fully healthy and green plant.
  • DI disease index
  • the term "quantitative trait locus” or “QTL” refers to a specific region on chromosomes, which harbors gene(s) controlling the trait.
  • the trait of particular interest is beet mild yellow virus (BMYV) resistance or resistance to BMYV.
  • a quantitative trait is a trait that varies in degree and which can be attributed to polygenic effects, i.e. a product of two or more genes.
  • locus refers to a certain place or position in the genome, e.g. on a chromosome or chromosome arm, which comprises one or more genetic determinants, for example one or several genes, contributing to a trait, such as a resistance to a disease.
  • a molecular marker refers to segment of nucleic acid that is inherited with a trait of interest.
  • a molecular marker may be a short DNA sequence, such as a sequence surrounding a single base-pair change, i.e. a single nucleotide polymorphism or SNP, or a long DNA sequence, such as microsatellites or Simple Sequence Repeats (SSRs).
  • SSRs Simple Sequence Repeats
  • the term “monomorph” as used herein describes the situation when a bi-allelic marker detects only one of two possible alleles in a given plant population. The lack of nucleotide variation for this position precludes linkage between a marker specific for this nucleotide and any trait that segregates in this population.
  • linked refers to a measurable probability that genes or markers located on a given chromosome are being passed on together to individuals in the next generation.
  • linked may refer to one or more genes or markers that are passed together with a gene with a probability greater than 0.5 (which is expected from independent assortment where markers/genes are located on different chromosomes).
  • interval marker refers to a marker that defines one of the termini of an interval (and is included in that interval). It will be clear that any of such intervals may comprise further markers. Two or more interval markers may be located at the respective termini of the region of interest (e.g. a QTL). For example, one interval marker may be located at the 5 ’ end of the QTL and one interval marker may be located at the 3’ end of the QTL.
  • interval refers to a continuous linear span of chromosomal DNA with termini defined by map position and/or markers.
  • a QTL positioned “between” two markers or “within an interval” is a QTL within a continuous linear span of chromosomal DNA that is flanked by two markers.
  • the “Refbeet 1.5 reference genome” as used herein refers to the assembled reference genome of the double-haploid KWS2320 line as published in Dohm JC, Minoche AE, Holtgrawe D, Capella-Gutierrez S, Zakrzewski F, Tafer H, Rupp O, Sorensen TR, Stracke R, Reinhardt R, Goesmann A, Kraft T, Schulz B, Stadler PF, Schmidt T, Gabaldon T, Lehrach H, Weisshaar B, Himmelbauer H. The genome of the recently domesticated crop plant sugar beet (Beta vulgaris). Nature. 2014 Jan 23;505(7484):546-9. doi: 10.1038/naturel2817.
  • Map positions in the reference genome refer to nucleotides of the respective chromosomes.
  • contig refers to a set of overlapping DNA segments that together represent a consensus region of DNA.
  • a contig refers to overlapping sequence data (reads); in top-down sequencing projects, contig refers to the overlapping clones that form a physical map of the genome that is used to guide sequencing and assembly. Contigs can thus refer both to overlapping DNA sequence and to overlapping physical segments (fragments) contained in clones depending on the context.
  • genomic DNA refers to extraction of the genomic DNA from cells obtained from one or several organisms.
  • KASP assay describes an assay for detection of SNP markers.
  • KASP genotyping assays are based on competitive allele-specific PCR and enable biallelic scoring of single nucleotide polymorphisms (SNPs) and insertions and deletions (Indels) at specific loci.
  • SNPs single nucleotide polymorphisms
  • Indels insertions and deletions
  • 70-100 base pairs upstream and 70-100 base pairs downstream of the SNP are selected and two allele -specific forward primers and one allele specific reverse primer is designed. See e.g. Allen et al. 2011, Plant Biotechnology J. 9, 1086-1099, especially pl 097- 1098 for KASP assay method.
  • SNP analysis refers to the detection of one or more specific SNPs within the DNA extracted from a sample from an individual organism or a population of organisms using known methods. The analysis typically includes comparison between individuals or populations or comparison with a reference sequence.
  • NGS next-generation sequencing
  • the term “Sanger sequencing”, also called “first-generation sequencing”, refers to a method of DNA sequencing based on the electrophoretic separation of chain-termination products produced in individual sequencing reactions.
  • nuclease based detection refers to a method of detection of DNA mismatches between two DNA fragments that differ in one nucleotide using a nuclease, e.g. Surveyor nuclease, that specifically detects mismatches in the DNA and then cleaves the DNA at the site of the mismatch.
  • a nuclease e.g. Surveyor nuclease
  • DNA chip technology refers to a collection of microscopic DNA molecules attached to a solid surface or beads.
  • DNA chip technology can for example be used for SNP detection, gene expression profiling or chromatin immunoprecipitation.
  • zygosity status refers to the type and number of allele present at a specific locus in the genome on a pair of homologous chromosomes. If both alleles of a diploid organism are the same, the organism is homozygous at that locus. If they are different, the organism is heterozygous at that locus. If one allele is missing, it is hemizygous, and, if both alleles are missing, it is nullizygous.
  • sequence Identity refers to the comparison of a first nucleic acid sequence to a second nucleic acid sequence, or a comparison of a first amino acid sequence to a second amino acid sequence and is calculated as a percentage based on the comparison. The result of this calculation can be described as “percent identical” or “percent ID.”
  • a sequence identity may be determined by a program, which produces an alignment, and calculates identity counting both mismatches at a single position and gaps at a single position as non-identical positions in final sequence identity calculation. The sequence identity is determined over the entire length of the first and second nucleic acid sequence.
  • sequence similarity or “similarity.” Means for making this adjustment are well-known to those of skill in the art. Typically this involves scoring a conservative substitution as a partial rather than a full mismatch, thereby increasing the percentage sequence identity.
  • a conservative substitution is given a score between zero and 1.
  • the scoring of conservative substitutions is calculated, e.g., according to the algorithm of Meyers and Miller, Computer Applic. Biol. Sci., 4: 11-17 (1988).
  • the comparison of sequences and determination of percent identity between two sequences can be accomplished using a mathematical algorithm.
  • An example of a local alignment algorithm utilized for the comparison of sequences is the NCBI Basic Local Alignment Search Tool (BLAST®) (Altschul et al. 1990 J. Mol. Biol.
  • NCBI National Center for Biotechnology Information
  • NLM National Library of Medicine
  • Another example of a mathematical algorithm utilized for the global comparison of sequences is the Clustal W and Clustal X (Larkin et al. 2007 Bioinformatics, 23, 2947-294, Clustal W and Clustal X version 2.0) as well as Clustal omega.
  • references to sequence identity used herein refer to the NCBI Basic Local Alignment Search Tool (BLAST®).
  • conferring resistance refers to a gene, nucleic acid, allele, marker, SNP, or coding variant resulting an increased resistance to a disease or pathogen.
  • transgene refers to at least one nucleic acid sequence that is taken from the genome of one organism, or produced synthetically, and which is then introduced into a host cell or organism or tissue of interest and which is subsequently integrated into the host's genome by means of transformation or transfection approaches.
  • transgenic refers to an organism carrying a transgene
  • transgenesis refers to the processes or methods of producing a transgenic organism.
  • genomic editing refers to strategies and techniques for the modification of any genetic information or genome by means of or involving a double stranded DNA break inducing enzyme or single-stranded DNA or RNA break inducing enzyme.
  • the terms comprise gene editing, but also the editing of regions other than gene encoding regions of a genome, such as intronic sequences, non-coding RNAs, miRNAs, sequences of regulatory elements like promoter, terminator, transcription activator binding sites, cis or trans acting elements.
  • the terms may comprise base editing for targeted replacement of single nucleobases. It can further comprise the editing of the nuclear genome as well as other genetic information, i.e. mitochondrial genome, chloroplast genome or an artificial genome or chromosome as well as miRNA, pre-mRNA or mRNA.
  • Targeted mutagenesis or “site-directed mutagenesis” as used herein refers to a method of making targeted, specific changes to a DNA sequence using methods of molecular biology.
  • Targeted mutagenesis includes, but is not limited to, CRISPR, TILLING, and TALEN methods.
  • non-targeted mutagenesis or “random mutagenesis” refers to a method wherein a mutagen or molecular biology method is used to introduce random changes into DNA. This can be achieved, for example, using UV radiation, mutagenic chemicals such as ethyl methane sulfonate (EMS) or nitrous acid, error prone PCR, DNA shuffling, insertion mutagenesis kits, or mutator strains with impaired DNA repair machinery.
  • EMS ethyl methane sulfonate
  • nitrous acid error prone PCR
  • DNA shuffling DNA shuffling
  • insertion mutagenesis kits or mutator strains with impaired DNA repair machinery.
  • harvested part refers to any picked or collected part of a crop plant.
  • saccharide refers to a product obtained from a sugar beet that can be used as a food product or in the production of biofuels.
  • derived products refers to any final product or by-product obtained from a beet plant.
  • resistance means the ability of a plant variety to restrict the infection, growth and/or development of a specified pest or pathogen (i.e., the host’s ability to limit pathogen multiplication) and the damage they cause when compared to susceptible plant varieties under similar environmental conditions and pest or pathogen pressure. Resistant varieties may exhibit some disease symptoms or damage under heavy pest or pathogen pressure. Following an infection by the virus, the viral load in the resistant variety (as measured for example in an ELISA assay or qPCR) is reduced in comparison to the susceptible variety.
  • the term “tolerance” or “tolerant” means the host’s ability to reduce the effect of infection on its fitness regardless of the level of pathogen multiplication.
  • the term is used herein to indicate a phenotype of a plant wherein disease symptoms remain reduced or absent upon exposure of said plant to an infective dosage of virus, whereby the presence of a systemic or local infection, virus multiplication, the presence of viral genomic sequences in cells of said plant and/or genomic integration thereof can be established. Tolerant plants are therefore carriers of the virus with less symptoms. Usually, viral sequences may be present or even multiply in plants without causing disease symptoms. Following an infection by the virus, the viral load in a tolerant variety (as measured for example in an ELISA assay or qPCR) is not substantially reduced.
  • progression refers to the incorporation of genetic material, such as alleles or genes, from one plant (e.g., a species or variety) into the gene pool of another.
  • human-induced random mutagenesis is intended to be understood in a conventional manner, i.e., to indicate an introduction of one or more mutations at random positions of the target sequence (i.e., as opposed to site-specific mutagenesis).
  • the random mutations are typically introduced by exposing plant, plant part, or plant cell to a mutagen and then screening for the presence of variants.
  • Mutagens for random mutagenesis are well known in the art, and include, for example, physical or chemical mutagenizing agent.
  • the term “mutagen” refers to a compound or process that results in the introduction of mutations in the plant genome.
  • mutagens include biological mutagens such as transposons, chemical mutagens such as N-nitroso-N-ethylurea, N-nitroso- N-methylurea, ethylmethane sulfonate (EMS), sodium azide, radiation, or other mutagens.
  • the radiation may be ultra-violet radiation, X-ray radiation, gamma radiation, alpha radiation, beta radiation, ion beams such as 4He ⁇ 2+> and H ⁇ +>, etc.
  • the additional trait or plant comprising said additional trait is not exclusively obtained by an essentially biological process.
  • the term "exclusively" in this context means that the method of establishing the additional trait and/or the plant comprising said additional trait is not entirely based on the crossing of whole genomes (i.e., an essentially biological process), but that the process contains - within the steps of sexually crossing and selecting - an additional step of a technical nature, which step by itself introduces a trait into the genome or modifies a trait in the genome of the plant produced, so that the introduction or modification of that trait is not the result of the mixing of the genes of the plants chosen for sexual crossing.
  • a trait or plant “established by a method such as genetic transformation, genome editing, targeted mutagenesis, and induced random mutagenesis” means a process which by itself introduces a trait into the genome or modifies a trait in the genome of the plant produced, so that the introduction or modification of that trait is not the result of the mixing of the genes of the plants chosen for sexual crossing.
  • Transformation of plant cells refers to process by which DNA is integrated into the genome of a plant cell.
  • the integration may be transient or stable.
  • “Stably” refers to the permanent, or non-transient retention and/or expression 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 may occur under natural or artificial conditions using various methods well known in the art.
  • Transformation may rely on any known method for the insertion of nucleic acid sequences into a prokaryotic or eukaryotic host cell, including Agrobacterium-mediated transformation protocols, viral infection, whiskers, electroporation, heat shock, lipofection, polyethylene glycol treatment, micro-injection, and particle bombardment.
  • Transgenic plant refers to genetically modified plant which contains at least one transgene.
  • Genus “Beta” belongs to the foxtail family (Amaranthaceae). The best known member is the common beet, Beta vulgaris, but several other species are recognized. These plants include plants of the species Beta macrocarpa, Beta vulgaris, Beta lomatogona, Beta macrorhiza, Beta coroffiflora, Beta trigyna and Beta nana.
  • a plant of the species Beta vulgaris is, in particular, a plant of the subspecies Beta vulgaris subsp. vulgaris. These include, for example, Beta vulgaris subsp. vulgaris var. altissima, Beta vulgaris ssp. vulgaris var. vulgaris (chard), Beta vulgaris ssp. vulgaris var. conditiva, Beta vulgaris ssp. vulgaris var. crassa/alba.
  • Beta vulgaris is a plant which is included in the subfamily Betoideae of the family Amaranthaceae. It is the economically most important crop of the large order Caryophyllales. It has several cultivar groups: the sugar beet, of greatest importance to produce table sugar; the root vegetable known as the beetroot or garden beet; the leaf vegetable known as chard or spinach beet; and Bruwurzel, which is a fodder crop. Three subspecies are typically recognized, but all cultivated beets fall into the subspecies Beta vulgaris subsp. vulgaris. The wild ancestor of the cultivated beets is the sea beet (Beta vulgaris subsp. maritima), and its Center of Origin lies in the Mediterranean region.
  • the beetroot is the taproot portion of a beet plant, usually known in Canada and the USA as beets, while the vegetable is referred to as beetroot in British English. It is also known as the table beet, garden beet, red beet, dinner beet or golden beet. It is one of several cultivar groups of Beta vulgaris grown for their edible taproots and leaves (called beet greens). These have been classified as B. vulgaris subsp. vulgaris ‘Conditiva’ Group.
  • Beta vulgaris (beet) is an herbaceous biennial or, rarely, perennial plant. Cultivated forms of beets are mostly biennial. The plant is usually erected with a long main root and a rosette of leaves growing on stems. The roots of cultivated forms are dark red, white, or yellow and moderately to strongly swollen and fleshy (subsp. vulgaris),' or brown, fibrous, sometimes swollen and woody in the wild subspecies. The stems grow erect or, in the wild forms, often procumbent; they are simple or branched in the upper part and their surface is ribbed and striate.
  • the basal leaves have a long petiole (which may be thickened and red, white, or yellow in some cultivars).
  • the simple leaf blade is oblanceolate to heart-shaped, dark green to dark red, slightly fleshy, usually with a prominent midrib, with entire or undulate margin, 5-20 cm long on wild plants (often much larger in cultivated plants about 20-40cm).
  • the upper leaves are smaller, their blades are rhombic to narrowly lanceolate.
  • the plant produces sessile green flowers and can reach 1-2 m in height. Beets are usually grown as annual plants and harvested after one growing season. The flowers are produced in dense spike-like, basally interrupted inflorescences. Very small flowers sit in one- to three- (rarely eight-) flowered glomerules in the axils of short bracts or in the upper half of the inflorescence without bracts.
  • the hermaphrodite flowers are urn-shaped, green or tinged reddish, and consist of five basally connate perianth segments (tepals), five stamens, and a semi -inferior ovary with two-three stigmas. The perianths of neighboring flowers are often fused.
  • the glomerules of flowers form connate hard clusters.
  • the fruit (utricle) is enclosed by the leathery and incurved perianth, and is immersed in the swollen, hardened perianth base.
  • the horizontal seed is lenticular, 2-3 mm, with a red-brown, shiny seed coat.
  • the seed contains an annular embryo and copious perisperm (feeding tissue). (Shultz, L. M 2003 Beta vulgaris. In: Flora of North America Editorial Committee (eds.): Flora of North America North of Mexico, Volume 4: “Magnoliophyta: Caryophyllidae”, part 1., Oxford University Press, New York, p.
  • the main vector aphids of yellowing are the green peach aphid (Myzus persicae) and the black bean aphid (Aphis fabae).
  • Myzus persicae the green peach aphid
  • Myzus persicae is the main vector for all aphid-transmitted virus species in sugar beet fields (Limburg, D.et al., (1997) Characteristics of beet yellows closterovirus transmission to sugar beets by Aphis fabae. Phytopathology, 87, 766-771; Schliephake, E., et al., (2000) Investigations on the vector transmission of the Beet mild yellowing virus (BMYV) and the Turnip yellows virus (TuYV).
  • BMYV Beet mild yellowing virus
  • TuYV Turnip yellows virus
  • BYV Beet Yellows Virus
  • Closterovirus characterized by a lemon-yellow coloration, which can subsequently cause small reddish necroses
  • BMYV Beet Mild Yellowing Virus
  • BWYV Beet Western Yellows Virus
  • BChV Beet Chlorosis Virus
  • BMYV, BChV, and BWYV are characterized by a more orange coloration, often followed by cryptogamic infestation (e.g. Altemaria) and premature leaf necrosis (see for example bisz.suedzucker.de/ssenschutz/blatt-krank phenomenon/viroese-vergilbung/ available on the worldwide web).
  • Beet yellows virus belongs to the genus Closterovirus in the family Closteroviridae . Infections with BYV lead to yellowish discoloration of the older leaves and subsequently reddish necrosis may occur (de Koeijer and van der Werf, 1999 Effects of Beet yellows virus and Beet mild yellowing virus on leaf area dynamics of sugar beet (Beta vulgaris L.). Field Crops Research, 61, 163-177).
  • BYV can be transmitted by more than 20 different aphid species in a semipersistent mode of transmission. In addition to M. persicae, Aphis fahae can contribute to virus transmission (Limburg et al., 1997). BYV virions move via the phloem but can also colonize mesophyll and epidermal cells and have been detected in plasmodesmata that connect the different cell types of the phloem (Dolja, V.V. (2003) Beet yellows virus', the importance of being different. Molecular Plant Pathology, 4, 91-98; Dolja, V.V. & Koonin, E.V. (2013) The closterovirus- derived gene expression and RNA interference vectors as tools for research and plant biotechnology. Frontiers in Microbiology, 4, 83).
  • Beet mild yellowing virus, Beet western yellows virus, and Beet chlorosis virus belong to the genus Polerovirus in the family Luteoviridae .
  • Beta species the viruses induce yellow to orange leaf discoloration, which may cause premature foliage death (Lewellen, R.T., et al. (1999) Reaction of sugar beet breeding lines and hybrids to beet chlorosis luteovirus. Journal of Sugar Beet Research, 36, 76).
  • Poleroviruses are persistently transmitted by their aphid vectors (Gray & Gildow, 2003 Luteovirus-aphid interactions. Annual Review of Phytopathology, 41, 539-566). Studies have shown that BMYV and BChV are efficiently transmitted by M. persicae (100%) and Macrosiphum euphorbiae (83%— 98%) (Kozlowska- Makulska et al., 2009).
  • BtMV Bactet mosaic virus belongs to the genus Potyvirus int the family Potyviridae. Symptoms of BtMV infection initially appear as yellowish speckles before the typical mosaic-like structures appear. In addition, the leaves are often malformed (Dunning & Byford, 1982 Pests, Diseases and Disorders of the Sugar Beet. Deleplanque: Brooms Bam Experimental Station). BtMV is transmitted via a nonpersistent mechanism (Gallet et al., 2018 Vector-transmission of plant viruses and constraints imposed by virus-vector interactions. Current Opinion in Virology, 33, 144-150). The main vectors for the transmission of BtMV in the field are M. persicae and A.
  • the vims can also be transmitted by many other species such as Myzus ascalonicus (Semal, J. (1956) Transmission of Beet mosaic virus from Stellaria media and Capsella hursa-pastoris by Myzus ascalonicus Doncaster. Nature, 178, 501-502), M. euphorhiae, Acyrthosiphon pisum, Metopolophium dirhodum, and Rhopalosiphum padi (Dusi & Peters, 1999).
  • the vector for BMYV vims is Myzus persicae.
  • BYV has primarily been controlled by controlling the insect vector through spay of insecticides.
  • climate change with warmer winters and the regulatory restriction on certain insecticides (especially neonicotinoid class) in the EU resulted in a significant increase of the problem.
  • the insecticides available to control the insect vectors have become very limited (currently only Tefluthrin products such as Force 20 CS).
  • Tefluthrin products such as Force 20 CS.
  • treatment with insecticides is not compatible with organic agriculture and/or the “Bio” label of resulting products.
  • Mamscha KWS has shown tolerance to BMYV. It is reported that under BMYV infection Mamscha KWS shows losses of 4% but still yields over 15t/ha more than the mean of KWS control varieties. Under BYV infection Mamscha KWS can show losses of 37% in yield, however it still yields 1 Ot/ha more than the mean of KW S UK non-tolerant commercial varieties under BYV infection. Mamscha KWS exhibits lower symptom expression - a greener canopy. However, in the absence of vims infection they do not reach the performance of non- tolerant varieties.
  • the genetic determinants for the trait was initially mapped using a backcrossed population in where the resistant germplasm was backcrossed with three different susceptible genotypes. From each cross, more than 180 BC1 -plants were analyzed using 95 AFLP -markers and phenotyped for BMYV resistance. The linkage between genetic region and trait was evaluated phenotypically in more than 2000 thousand individual plants to ensure data of required quality. The genetic determinants were then identified using statistical methods (t-test and power analysis). Fine mapping was then carried out by selecting one of the previously mentioned backcrosses and creating four different F2 populations by selfing four BC1 -plants.
  • an additional mapping population was established by crossing the resistant germplasm with an unrelated susceptible germplasm to form an Fl -population. From this initial cross, using a single-seed lineage method, an F2 population consisting of more than 1000 individuals in which all mapped genetic determinants were segregating was established. From this populations, more than 450 individuals were selected and selfed to create an F2S1 population. The individuals in this population was then further selfed to create an F2S2 population. Each parental plant in the F2S1 generation was genotyped using more than 21 000 SNP-markers. The genetic determinants were then identified using statistical genetic analysis tools for inheritance based trait mapping as well as multiple marker association models. The position and extent of the genetic determiats were determined using both maximum likelihood single point estimation (MLSPE) as well as through 95% Bayesian interval mapping. A total of 27 trait specific markers were developed.
  • MLSPE maximum likelihood single point estimation
  • QTL1 is located on chromosome 1 and genetically linked to at least one marker loci as follows: PBV504245 is a single nucleotide polymorphism (SNP) corresponding to position 1050427 of chromosome 1 of the Refbeet 1.5 reference genome, wherein said nucleotide is G or C, with G being the nucleotide for the resistant line; PBV125552 is a SNP corresponding to position 3069972 of chromosome 1 of the Refbeet 1.5 reference genome, wherein said nucleotide is A or G, with A being the nucleotide for the resistant line; PBV291217 is a SNP corresponding to position 4315236 of chromosome 1 of the Refbeet 1.5 reference genome, wherein said nucleotide is A or G,
  • QTL2 is located on chromosome 4 and genetically linked to at least one marker loci as follows: selected from the group consisting of PBV577030 is a SNP corresponding to position 5742753 of chromosome 4 of the Refbeet 1.5 reference genome, wherein said nucleotide is G or A, with G being the nucleotide for the resistant line; PBV562771 is a SNP corresponding to position 9767085 of chromosome 4 of the Refbeet 1.5 reference genome, wherein said nucleotide is A or G, with A being the nucleotide for the resistant line; PBV573138 is a SNP corresponding to position 10271308 of chromosome 4 of the Refbeet 1.5 reference genome, wherein said nucleotide is G or A, with G being the nucleotide for the resistant line; PBV273480 is a SNP corresponding to position 46718199 of chromosome 4 of the Refbeet 1.5 reference
  • QTL3 is located on chromosome 8 and genetically linked to at least one marker loci as follows: PBV148969 is a SNP corresponding to position 436005 of chromosome 8 of the Refbeet 1.5 reference genome, wherein said nucleotide is G or A, with G being the nucleotide for the resistant line; PBV331644 is a SNP corresponding to position 1366650 of chromosome 8 of the Refbeet 1.5 reference genome, wherein said nucleotide is A or G, with A being the nucleotide for the resistant line; PBV882826 is a SNP corresponding to position 7979633 of chromosome 8 of the Refbeet 1.5 reference genome, wherein said nucleotide is G or A, with G being the nucleotide for the resistant line; PBV374371 is a SNP corresponding to position 8822544 of chromosome 8 of the Refbeet 1.5 reference genome, wherein said nucleot
  • QTL4 is located on chromosome 2 and is genetically linked to at least one marker loci as follows: GRKPBV98281 is a SNP corresponding to position 8040503 of chromosome 2 of the Refbeet 1.5 reference genome, wherein said nucleotide is G or A, with A being the nucleotide for the resistant line; GRKPBV627372 is a SNP corresponding to position 9433027 of chromosome 2 of the Refbeet 1.5 reference genome, wherein said nucleotide is C or A, with A being the nucleotide for the resistant line; GRKPBV672273 is a SNP corresponding to position 12212298 of chromosome 2 of the Refbeet 1.5 reference genome, wherein said nucleotide is G or A, with A being the nucleotide for the resistant line; GRKPBV627178 is a SNP corresponding to position 18410288 of chromosome 2 of the Re
  • QTL1, QTL2, QTL 3, and QTL 4 are obtainable from a Beta plant, such as Beta vulgaris subsp. vulgaris line 21000003-71, representative seed of which has been deposited under Deposit Number NCIMB 44107.
  • the present invention provides plants, plant parts, and plant cells of the genus Beta resistant to beet yellowing disease(s) caused by Beet yellowing virus(s).
  • the inventors identified resistant germplasm and mapped the resistant trait to four quantitative trait loci, QTL1, QTL2, QTL3, and QTL4. They further identified multiple markers for each QTL and have crossed the BMYV resistance trait into Beta genetic backgrounds, thus obtaining resistant Beta plants, especially, sugar beet (Beta vulgaris subsp. vulgaris) plant.
  • the present disclosure provides a Beta vulgaris subsp. vulgaris plant, plant part, or plant cell resistant to BMYV, wherein said plant, plant part, or plant cell comprises at least one of QTL1, QTL2, QTL3, and QTL 4.
  • the QTL1 on chromosome 1 is genetically linked to at least one of markers PBV504245, PBV125552, PBV291217, PBV369685, PBV261963, PBV757437, PBV638722, and EPBV6296.
  • the QTL2 on chromosome 4 is genetically linked to at least one of markers PBV577030, PBV562771, PBV573138, PBV273480, and PBV82I05I.
  • the QTL3 on chromosome 8 is genetically linked to at least one of markers PBV148969, PBV331644, PBV882826, PBV374371, PBV757436, PBV738292, PBV738293, and EPBV7349.
  • the QTL4 on chromosome 2 is genetically linked to at least one of markers GRKPBV98281, GRKPBV627372, GRKPBV672273, GRKPBV627178, GRKPBV116271, and EPBV8821.
  • said QTLs conferring resistance to BMYV are located from about 20 cM to about 30 cM of said markers.
  • the disclosure relates to a Beta vulgaris subsp. vulgaris plant, plant part, or plant cell heterozygous for QTL1, QTL2, QTL3 and QTL4.
  • the disclosure relates to a. Beta vulgaris subsp. vulgaris plant, plant part, or plant cell homozygous for QTL1, QTL2, QTL3, and QTL4.
  • the disclosure relates to a Beta vulgaris subsp. vulgaris plant, plant part, or plant cell heterozygous for at least one of QTL1, QTL2, QTL3, and QTL4. In some embodiments, the disclosure relates to a Beta vulgaris subsp. vulgaris plant, plant part, or plant cell homozygous for at least one of QTL1, QTL2, QTL3, and QTL4.
  • the disclosure relates to a Beta vulgaris subsp. vulgaris plant, plant part, or plant cell heterozygous for QTL1 and homozygous for at least one of QTL2, QTL3, and QTL4.
  • the disclosure relates to a Beta vulgaris subsp. vulgaris plant, plant part, or plant cell heterozygous for QTL2 and homozygous for at least one of QTL1, QTL3, and QTL4.
  • the disclosure relates to a Beta vulgaris subsp. vulgaris plant, plant part, or plant cell heterozygous for QTL3 and homozygous for at least one of QTL1, QTL2, and QTL4.
  • the disclosure relates to a Beta vulgaris subsp. vulgaris plant, plant part, or plant cell heterozygous for QTL4 and homozygous for at least one of QTL1, QTL2, and QTL3.
  • the disclosure relates to a Beta vulgaris subsp. vulgaris plant, plant part, or plant cell homozygous for QTL1 and heterozygous for at least one of QTL2, QTL3, and QTL4.
  • the disclosure relates to a Beta vulgaris subsp. vulgaris plant, plant part, or plant cell homozygous for QTL2 and heterozygous for at least one of QTL1, QTL3, and QTL4.
  • the disclosure relates to a Beta vulgaris subsp. vulgaris plant, plant part, or plant cell homozygous for QTL3 and heterozygous for at least one of QTL1, QTL2, and QTL4.
  • the disclosure relates to a Beta vulgaris subsp. vulgaris plant, plant part, or plant cell homozygous for QTL4 and heterozygous for at least one of QTL1, QTL2, and QTL3.
  • the development of plants resistant against Beet Yellows Virus is taught herein.
  • the present disclosure provides a genetic resistance solution to the problem.
  • the resistance works primarily against Beet Mild Yellow Virus and partially for others some Yellow Viruses. It prevents yield loss from viral infection.
  • the trait is differentiated from the Maruscha and Novalina varieties in that the resistance effectively and significantly lowers the viral load as demonstrated by ELISA assay. This results in a strong suppression of the visible symptoms and protects yield.
  • the trait is dominant (or at least partially dominant) and can confer resistance in heterozygous or homozygous form. This enables creating multiple hybrids without major efforts in introgression.
  • the disclosure is applicable to sugar beet, fodder beet, but potentially also to vegetable beet crops (red beet etc.). It is especially beneficial for varieties which are targeting the bio- organic sector, where chemical insecticides cannot be used.
  • seeds obtained from the yellowing -virus resistant beet plants are planted in areas where the use of insecticides to control the vector for BMYV transmission is restricted, prohibited, or not desired.
  • plants germinated from the seeds are grown under the controlled conditions or environments without the use of insecticides.
  • the plants produce roots and/or beets, which can be used to produce sugar that is qualified for the organic market.
  • organic or “organically grown” refers to plant or food grown and processed using no synthetic fertilizers, insecticides or pesticides. Pesticides derived from natural sources (such as biological pesticides) may be used in producing organically grown plant or food. As a result, commodity products from organic or organically grown plants is free of the effect of synthetic fertilizers, pesticides, or insecticides, fungicides, nematicides, etc.
  • the pesticides used to control the disease are not allowed in organic farming.
  • organic cultivation of sugar beet in virus yellows infested regions will require a strong genetic resistance as the currently used pesticides are not allowed or applied in organic farming.
  • a seed of the yellowing virus resistant plant is technically treated, whereby the technical treatment is selected from the group consisting of polishing, pelleting, incrustation, and coloring.
  • the technical treatment does not include a pesticide, an insecticide, a fungicide, or a nematicide.
  • the goal of beet breeding is to develop new, unique and superior beet cultivar and hybrids.
  • the breeder initially selects and crosses two or more parental lines, followed by repeated selfing and selection, producing many new genetic combinations.
  • Another method used to develop new, unique and superior beet cultivars occurs when the breeder selects and crosses two or more parental lines followed by haploid induction and chromosome doubling that result in the development of dihaploid cultivars.
  • the breeder can theoretically generate billions of different genetic combinations via crossing, selfing and mutations and the same is true for the utilization of the dihaploid breeding method.
  • Pedigree breeding and recurrent selection breeding methods are used to develop cultivars from breeding populations. Breeding programs combine desirable traits from two or more cultivars or various broad-based sources into breeding pools from which cultivars are developed by selfing and selection of desired phenotypes or through the dihaploid breeding method followed by the selection of desired phenotypes. The new cultivars are evaluated to determine which have commercial potential.
  • Choice of breeding or selection methods depends on the mode of plant reproduction, the heritability of the trait(s) being improved, and the type of cultivar used commercially (e.g., Fi hybrid cultivar, pureline cultivar, etc.). For highly heritable traits, a choice of superior individual plants evaluated at a single location will be effective, whereas for traits with low heritability, selection should be based on mean values obtained from replicated evaluations of families of related plants.
  • Popular selection methods commonly include pedigree selection, modified pedigree selection, mass selection, recurrent selection, and backcross breeding.
  • the plants of the present disclosure can be used to produce new plant varieties.
  • the plants are used to develop new, unique and superior varieties or hybrids with desired phenotypes.
  • plant breeding techniques comprises all of the plant breeding techniques disclosed in this section of the application, and well known to persons having skill in the art.
  • plant breeding methods encompass the application of recurrent selection, mass selection, hybridization, open-pollination, backcrossing, pedigree breeding, marker assisted selection breeding, mutation breeding, double haploids and chromosome doubling, gene editing, and combinations thereof.
  • selection methods e.g., molecular marker assisted selection
  • breeding methods can be combined with breeding methods to accelerate the process. Additional breeding methods have been known to one of ordinary skill in the art, e.g., methods discussed in Chahal and Gosal (Principles and procedures of plant breeding: biotechnological and conventional approaches, CRC Press, 2002, ISBN 084931321X, 9780849313219), Taji et al.
  • Beta vulgaris sucgar beet genome has been sequenced recently (Dohm et. al. (2014) The genome of the recently domesticated crop plant sugar beet (Beta vulgaris). Nature. 2014 Jan 23;505(7484):546- 9; Minoche et. al.
  • molecular markers are designed and made, based on the genome of the plants of the present application.
  • the molecular markers are selected from Isozyme Electrophoresis, Restriction Fragment Length Polymorphisms (RFLPs), Randomly Amplified Polymorphic DNAs (RAPDs), Arbitrarily Primed Polymerase Chain Reaction (AP-PCR), DNA Amplification Fingerprinting (DAF), Sequence Characterized Amplified Regions (SCARs). Amplified Fragment Length Polymorphisms (AFLPs), and Simple Sequence Repeats (SSRs) which are also referred to as Microsatellites, etc.
  • RFLPs Restriction Fragment Length Polymorphisms
  • RAPDs Randomly Amplified Polymorphic DNAs
  • AP-PCR Arbitrarily Primed Polymerase Chain Reaction
  • DAF DNA Amplification Fingerprinting
  • SCARs Sequence Characterized Amplified Regions
  • AFLPs Amplified
  • the molecular markers can be used in molecular marker assisted breeding.
  • the molecular markers can be utilized to monitor the transfer of the genetic material.
  • the transferred genetic material is a gene of interest, such as genes that contribute to one or more favorable agronomic phenotypes when expressed in a plant cell, a plant part, or a plant.
  • Classical breeding methods can be included in the present disclosure to introduce one or more yellowing virus-resistant traits of the present disclosure into other plant varieties, or other close-related species that are compatible to be crossed with the transgenic plant.
  • the yellowing virus-resistant traits can be resistance against BMYV, BYV, BChV, BtMV, and BWYW, as disclosed in this application.
  • said method comprises (i) crossing any one of the plants of the present disclosure resistant to yellowing virus (such as BMYV, BYV, BChV, BtMV, and BWYW) as a donor to a recipient plant line to create a Fl population; (ii) selecting offspring resistant to the yellowing virus.
  • the offspring can be further selected by testing or diagnosing the presence of QTLs taught herein.
  • complete chromosomes or desired QTLs of the donor plant are transferred.
  • the yellowing virus-resistant plant with at least one of QTLs of the present disclosure can serve as a male or female parent in a cross pollination to produce offspring plants, wherein by receiving the QTLs from the donor plant, the offspring plants have at least one of the QTLs.
  • protoplast fusion can also be used for the transfer of the transgene from a donor plant to a recipient plant.
  • Protoplast fusion is an induced or spontaneous union, such as a somatic hybridization, between two or more protoplasts (cells in which the cell walls are removed by enzymatic treatment) to produce a single bi- or multi-nucleate cell.
  • the fused cell that may even be obtained with plant species that cannot be interbred in nature is tissue cultured into a hybrid plant exhibiting the desirable combination of traits. More specifically, a first protoplast can be obtained from a plant resistant to yellowing virus.
  • a second protoplast can be obtained from a second plant line, optionally from another plant species or variety, preferably from the same plant species or variety, that comprises commercially desirable characteristics, such as, but not limited to disease resistance, insect resistance, valuable grain characteristics (e.g., increased seed weight and/or seed size) etc.
  • the protoplasts are then fused using traditional protoplast fusion procedures, which are known in the art to produce the cross.
  • embryo rescue may be employed in the transfer of the QTLs from a donor plant to a recipient plant.
  • Embryo rescue can be used as a procedure to isolate embryos from crosses wherein plants fail to produce viable seed.
  • the fertilized ovary or immature seed of a plant is tissue cultured to create new plants (see Pierik, 1999, In vitro culture of higher plants, Springer, ISBN 079235267x, 9780792352679, which is incorporated herein by reference in its entirety).
  • the recipient plant is an elite line having one or more certain desired traits.
  • desired traits include but are not limited to those that result in increased biomass production, production of specific chemicals, increased seed production, improved plant material quality, increased seed oil content, etc. Additional examples of desired traits include pest resistance, vigor, development time (time to harvest), enhanced nutrient content, novel growth patterns, aromas or colors, salt, heat, drought and cold tolerance, and the like.
  • Desired traits also include selectable marker genes (e.g., genes encoding herbicide or antibiotic resistance used only to facilitate detection or selection of transformed cells), hormone biosynthesis genes leading to the production of a plant hormone (e.g., auxins, gibberellins, cytokinins, abscisic acid and ethylene that are used only for selection), or reporter genes (e.g. luciferase, [3-glucuronidase, chloramphenicol acetyl transferase (CAT, etc.).
  • the recipient plant can also be a plant with preferred chemical compositions, e.g., compositions preferred for medical use or industrial applications.
  • Open-Pollinated Populations The improvement of open-pollinated populations of such crops as beet, rye, many maizes and sugar beets, herbage grasses, legumes such as alfalfa and clover, and tropical tree crops such as cacao, coconuts, oil palm and some rubber, depends essentially upon changing gene-frequencies towards fixation of favorable alleles while maintaining a high (but far from maximal) degree of heterozygosity. Uniformity in such populations is impossible and trueness-to-type in an open-pollinated variety is a statistical feature of the population as a whole, not a characteristic of individual plants. Thus, the heterogeneity of open-pollinated populations contrasts with the homogeneity (or virtually so) of inbred lines, clones and hybrids.
  • Population improvement methods fall naturally into two groups, those based on purely phenotypic selection, normally called mass selection, and those based on selection with progeny testing.
  • Interpopulation improvement utilizes the concept of open breeding populations; allowing genes to flow from one population to another. Plants in one population (cultivar, strain, ecotype, or any germplasm source) are crossed either naturally (e.g., by wind) or by hand or by bees (commonly Apis mellifera L. or Megachile rotundata F. ) with plants from other populations. Selection is applied to improve one (or sometimes both) population(s) by isolating plants with desirable traits from both sources.
  • Mass Selection In mass selection, desirable individual plants are chosen, harvested, and the seed composited without progeny testing to produce the following generation. Since selection is based on the maternal parent only, and there is no control over pollination, mass selection amounts to a form of random mating with selection. As stated herein, the purpose of mass selection is to increase the proportion of superior genotypes in the population.
  • Mutation breeding is another method of introducing new traits into the beet plants of the present disclosure. Mutations that occur spontaneously or are artificially induced can be useful sources of variability for a plant breeder. The goal of artificial mutagenesis is to increase the rate of mutation for a desired characteristic.
  • Mutation rates can be increased by many different means including temperature, long-term seed storage, tissue culture conditions, radiation; such as X-rays, Gamma rays (e.g., cobalt 60 or cesium 137), neutrons, (product of nuclear fission by uranium 235 in an atomic reactor), Beta radiation (emitted from radioisotopes such as phosphorus 32 or carbon 14), or ultraviolet radiation (preferably from 2500 to 2900 nm), or chemical mutagens (such as base analogues (5 -bromo-uracil)), related compounds (8-ethoxy caffeine), antibiotics (streptonigrin), alkylating agents (sulfur mustards, nitrogen mustards, epoxides, ethyleneamines, sulfates, sulfonates, sulfones, lactones), azide, hydroxylamine, nitrous acid, or acridines.
  • radiation such as X-rays, Gamma rays (e.g.,
  • Double Haploids and Chromosome Doubling One way to obtain homozygous plants without the need to cross two parental lines followed by a long selection of the segregating progeny, and/or multiple backcrossing is to produce haploids and then double the chromosomes to form doubled haploids.
  • Haploid plants can occur spontaneously or may be artificially induced via chemical treatments or by crossing plants with inducer lines (Seymour et al. 2012, PNAS vol 109, pg 4227-4232; Zhang et al., 2008 Plant Cell Rep. Dec 27(12) 1851-60).
  • the production of haploid progeny can occur via a variety of mechanisms which can affect the distribution of chromosomes during gamete formation.
  • Additional methods include, but are not limited to, expression vectors introduced into plant tissues using a direct gene transfer method, such as microprojectile-mediated delivery, DNA injection, electroporation, and the like. Additionally, expression vectors are introduced into plant tissues by using either microprojectile-mediated delivery with a biolistic device or by using Agrobacterium-mediated transformation. Transformant plants obtained with the protoplasm of the subject beet plants are intended to be within the scope of the embodiments of the application.
  • breeding and selection schemes of the present disclosure can include crosses with plant lines that have undergone genome editing.
  • the breeding and selection methods of the present disclosure are compatible with plants that have been modified using any gene and/or genome editing tool, including, but not limited to: ZFNs, TALENS, CRISPR, and Mega nuclease technologies.
  • any gene and/or genome editing tool including, but not limited to: ZFNs, TALENS, CRISPR, and Mega nuclease technologies.
  • persons having skill in the art will recognize that the breeding methods of the present disclosure are compatible with many other gene editing technologies.
  • the present disclosure teaches gene-editing technologies can be applied for a single locus conversion, for example, conferring beet plant with virus resistance.
  • the present disclosure teaches that the single locus conversion is an artificially mutated gene or nucleotide sequence that has been modified through the use of breeding techniques taught herein.
  • ZFN-3 Genes encoding ZFNs are delivered to plant cells along with a stretch of DNA which can be several kilo base pairs long and the ends of which are homologous to the DNA sequences flanking the cleavage site. As a result, the DNA stretch is inserted into the plant genome in a site-specific manner.
  • the breeding and selection methods of the present disclosure are compatible with plants that have been modified through Transcription activator-like (TAL) effector nucleases (TALENs).
  • TALENS are polypeptides with repeat polypeptide arms capable of recognizing and binding to specific nucleic acid regions. By engineering the polypeptide arms to recognize selected target sequences, the TAL nucleases can be used to direct double stranded DNA breaks to specific genomic regions. These breaks can then be repaired via recombination to edit, delete, insert, or otherwise modify the DNA of a host organism.
  • TALENSs are used alone for gene editing (e.g., for the deletion or disruption of a gene).
  • the breeding and selection methods of the present disclosure are compatible with plants that have been modified through meganucleases.
  • meganucleases are engineered endonucleases capable of targeting selected DNA sequences and inducing DNA breaks.
  • new meganucleases targeting specific regions are developed through recombinant techniques which combine the DNA binding motifs from various other identified nucleases.
  • new meganucleases are created through semi-rational mutational analysis, which attempts to modify the structure of existing binding domains to obtain specificity for additional sequences.
  • breeding schemes of the present application can include crosses between donor and recipient plants.
  • said donor plants contain a gene or genes of interest which may confer the plant with a desirable phenotype.
  • the recipient line can be an elite line or cultivar having certain favorable traits for commercial production.
  • the elite line may contain other genes that also impart said line with the desired phenotype.
  • the donor and recipient plant may create a progeny plant with combined desirable loci which may provide quantitatively additive effect of a particular characteristic. In that case, QTL mapping can be involved to facilitate the breeding process.
  • One or more such QTLs associated with a desirable trait in a donor plant can be transferred to a recipient plant to incorporate the desirable trait into progeny plants by transferring and/or breeding methods.
  • Isogenic lines in which favorable QTL alleles have been fixed can be generated by systematic backcrossing. These isogenic lines may be referred to as near-isogenic lines (NILs), introgression lines (ILs), backcross inbred lines (BILs), backcross recombinant inbred lines (BCRIL), recombinant chromosome substitution lines (RCSLs), chromosome segment substitution lines (CSSLs), and stepped aligned inbred recombinant strains (STAIRSs).
  • NILs near-isogenic lines
  • ILs introgression lines
  • BILs backcross inbred lines
  • BCRIL backcross recombinant inbred lines
  • RCSLs recombinant chromosome substitution lines
  • CSSLs chromosome segment substitution lines
  • STAIRSs stepped aligned inbred recombinant strains
  • the plants are selected on the basis of one or more phenotypic traits.
  • phenotypic traits include any observable characteristic of the plant, including for example growth rate, vigor, plant health, maturity, branching, plant height, leaf coverage, weight, total yield, color, taste, sugar levels, aroma, changes in the production of one or more compounds by the plant (including for example, metabolites, proteins, drugs, carbohydrates, oils, and any other compounds).
  • a most difficult task is the identification of individuals that are genetically superior, because for most traits the true genotypic value is masked by other confounding plant traits or environmental factors.
  • One method of identifying a superior plant is to observe its performance relative to other experimental plants and to a widely grown standard cultivar. If a single observation is inconclusive, replicated observations provide a better estimate of its genetic worth.
  • plants may be selected based on the absence, suppression or inhibition of a certain feature or trait (such as an undesirable feature or trait) as opposed to the presence of a certain feature or trait (such as a desirable feature or trait).
  • genotypic information for example, including the pattern of plant gene expression, genotype, or presence of genetic markers.
  • the one or more marker may already be known and/or associated with a particular characteristic of a plant; for example, a marker or markers may be associated with an increased growth rate or metabolite profile.
  • This information could be used in combination with assessment based on other characteristics in a method of the disclosure to select for a combination of different plant characteristics that may be desirable.
  • Such techniques may be used to identify novel quantitative trait loci (QTLs).
  • plants may be selected based on growth rate, size (including but not limited to weight, height, leaf size, stem size, branching pattern, or the size of any part of the plant), general health, survival, tolerance to adverse physical environments and/or any other characteristic, as described herein before.
  • Non-limiting examples include selecting plants based on: speed of seed germination; quantity of biomass produced; increased root, and/or leaf/ shoot growth that leads to an increased yield (herbage or grain or fiber or oil) or biomass production; effects on plant growth that results in an increased root and/or seed yield for a crop; effects on plant growth which result in an increased head yield; effects on plant growth that lead to an increased resistance or tolerance to disease including fungal, viral or bacterial diseases, to mycoplasma, or to pests such as insects, mites or nematodes in which damage is measured by decreased foliar symptoms such as the incidence of bacterial or fungal lesions, or area of damaged foliage or reduction in the numbers of nematode cysts or galls on plant roots, or improvements in plant yield in the presence of such plant pests and diseases; effects on plant growth that lead to increased metabolite yields; effects on plant growth that lead to improved aesthetic appeal which may be particularly important in plants grown for their form, color or taste, for example the color intensity of beet leaves/
  • Selection of plants based on phenotypic or genotypic information may be performed using techniques such as, but not limited to: high through-put screening of chemical components of plant origin, sequencing techniques including high through-put sequencing of genetic material, differential display techniques (including DDRT-PCR, and DD-PCR), nucleic acid microarray techniques, RNA-seq (Transcriptome Sequencing), qPCR (quantitative real time PCR) and RT-qPCR (reverse transcriptase quantitative real time PCR).
  • high through-put screening of chemical components of plant origin sequencing techniques including high through-put sequencing of genetic material
  • differential display techniques including DDRT-PCR, and DD-PCR
  • nucleic acid microarray techniques including DDRT-PCR, and DD-PCR
  • RNA-seq Transcriptome Sequencing
  • qPCR Quantitative real time PCR
  • RT-qPCR reverse transcriptase quantitative real time PCR
  • the evaluating step of a plant breeding program involves the identification of desirable traits in progeny plants.
  • Progeny plants can be grown in, or exposed to conditions designed to emphasize a particular trait (e.g. drought conditions for drought tolerance, lower temperatures for freezing tolerant traits).
  • Progeny plants with the highest scores for a particular trait may be used for subsequent breeding steps.
  • plants selected from the evaluation step can exhibit a 1%, 5%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 100%, 120% or more improvement in a particular plant trait compared to a control plant.
  • the evaluating step of plant breeding comprises one or more molecular biological tests for genes or other markers.
  • the molecular biological test can involve probe hybridization and/or amplification of nucleic acid (e.g., measuring nucleic acid density by Northern or Southern hybridization, PCR) and/or immunological detection (e.g., measuring protein density, such as precipitation and agglutination tests, ELISA (e.g., Lateral Flow test or DAS-ELISA), Western blot, immune labeling, immunosorbent electron microscopy (ISEM), and/or dot blot).
  • nucleic acid e.g., measuring nucleic acid density by Northern or Southern hybridization, PCR
  • immunological detection e.g., measuring protein density, such as precipitation and agglutination tests, ELISA (e.g., Lateral Flow test or DAS-ELISA), Western blot, immune labeling, immunosorbent electron microscopy (ISEM), and/or dot blot.
  • nucleic acid hybridization an amplification of nucleic acid (e.g., PCR, RT-PCR) or an immunological detection (e.g., precipitation and agglutination tests, ELISA (e.g., Lateral Flow test or DAS-ELISA), Western blot, RIA, immunogold or immunofluorescent labeling, immunosorbent electron microscopy (ISEM), and/or dot blot tests) are performed as described elsewhere herein and well-known by one skilled in the art.
  • an amplification of nucleic acid e.g., PCR, RT-PCR
  • immunological detection e.g., precipitation and agglutination tests
  • ELISA e.g., Lateral Flow test or DAS-ELISA
  • Western blot RIA
  • immunogold or immunofluorescent labeling e.g., immunosorbent electron microscopy (ISEM), and/or dot blot tests
  • the evaluating step comprises PCR (semi-quantitative or quantitative), wherein primers are used to amplify one or more nucleic acid sequences of a desirable gene, or a nucleic acid associated with said gene, or QTL or a desirable trait (e.g., a co-segregating nucleic acid, or other marker).
  • primers are used to amplify one or more nucleic acid sequences of a desirable gene, or a nucleic acid associated with said gene, or QTL or a desirable trait (e.g., a co-segregating nucleic acid, or other marker).
  • the evaluating step comprises immunological detection (e.g., precipitation and agglutination tests, ELISA (e.g., Lateral Flow test or DAS-ELISA), Western blot, RIA, immuno labeling (gold, fluorescent, or other detectable marker), immunosorbent electron microscopy (ISEM), and/or dot blot), wherein one or more gene or marker-specific antibodies are used to detect one or more desirable proteins.
  • said specific antibody is selected from the group consisting of polyclonal antibodies, monoclonal antibodies, antibody fragments, and combination thereof.
  • RT-PCR Reverse Transcription Polymerase Chain Reaction
  • PCR polymerase chain reaction
  • amplification a laboratory technique commonly used in molecular biology to generate many copies of a DNA sequence.
  • RNA strand is first reverse transcribed into its DNA complement (complementary DNA, or cDNA) using the enzyme reverse transcriptase, and the resulting cDNA is amplified using traditional or real-time PCR.
  • Real time RT-PCR provides a method where the amplicons can be visualized as the amplification progresses using a fluorescent reporter molecule.
  • fluorescent reporters There are three major kinds of fluorescent reporters used in real time RT-PCR, general non-specific DNA Binding Dyes such as SYBR Green I, TaqMan Probes and Molecular Beacons (including Scorpions).
  • nucleic acid detection can include next-generation sequencing methods such as DNA SEQ or RNA SEQ using any known sequencing platform including, but not limited to: Roche 454, Solexa Genome Analyzer, AB SOLiD, Illumina GA/HiSeq, Ion PGM, Mi Seq, among others (Liu et al,. 2012 Journal of Biomedicine and Biotechnology Volume 2012 ID 251364; Franca et al., 2002 Quarterly Reviews of Biophysics 35 pg 169-200; Mardis 2008 Genomics and Human Genetics vol 9 pg 387-402).
  • next-generation sequencing methods such as DNA SEQ or RNA SEQ using any known sequencing platform including, but not limited to: Roche 454, Solexa Genome Analyzer, AB SOLiD, Illumina GA/HiSeq, Ion PGM, Mi Seq, among others
  • nucleic acids may be detected with other high-throughput hybridization technologies including microarrays, gene chips, LNA probes, nanoStrings, and fluorescence polarization detection, among others.
  • detection of markers can be achieved at an early stage of plant growth by harvesting a small tissue sample (e.g., branch, or leaf disk). This approach is preferable when working with large populations as it allows breeders to weed out undesirable progeny at an early stage and conserve growth space and resources for progeny which show more promise.
  • the detection of markers is automated, such that the detection and storage of marker data is handled by a machine. Recent advances in robotics have also led to full-service analysis tools capable of handling nucleic acid/protein marker extractions, detection, storage and analysis.
  • Plant breeding begins with the analysis and definition of problems and weaknesses of the current germplasm, the establishment of program goals, and the definition of specific breeding objectives.
  • the next step is selection of germplasm that possesses the traits to meet the program goals.
  • the goal is to combine in a single variety or hybrid an improved combination of desirable traits from the parental germplasm.
  • these important traits may include higher yield of roots or leaves, improved leaf color, consistent leaf color at the juvenile stage in warm as well as cool conditions, agronomic quality such as sugar level, root size, root color or texture, root firmness, resistance to diseases and insects, adaptability for soil and climate conditions, harvest flexibility, tolerance to drought and heat, improved post-harvest shelf-life of the leaves or postharvest quality of roots, improved standing ability in the field, improved uniformity of leaves or roots, improved bolting tolerance in seed production, improved seedling vigor, and the like.
  • agronomic quality such as sugar level, root size, root color or texture, root firmness, resistance to diseases and insects, adaptability for soil and climate conditions, harvest flexibility, tolerance to drought and heat, improved post-harvest shelf-life of the leaves or postharvest quality of roots, improved standing ability in the field, improved uniformity of leaves or roots, improved bolting tolerance in seed production, improved seedling vigor, and the like.
  • receivers of the desired traits can be any susceptible individual belonging to a species crossable with sugar beet such as members of the Beta vulgaris ssp. vulgaris group which includes crops such as sugarbeet, chard, beetroot and mangold but also closely related species such as Beta vulgaris ssp. maritima or any other member of the Betoideae family able to produce viable offspring when crossed with Beta vulgaris ssp. vulgaris.
  • a species crossable with sugar beet such as members of the Beta vulgaris ssp. vulgaris group which includes crops such as sugarbeet, chard, beetroot and mangold but also closely related species such as Beta vulgaris ssp. maritima or any other member of the Betoideae family able to produce viable offspring when crossed with Beta vulgaris ssp. vulgaris.
  • a plant of the present disclosure can be crossed with a .Be genus plant that is crossable (e.g., sexually compatible).
  • the genus Beta to which B. vulgaris belongs, is comprised of 15 recognized species which are divided into four sections: Beta, Corollinae, Procumbentes and Nanae.
  • B. vulgaris and specific members within the Beta section i.e. fodder beet, red beet, leaf beet, Swiss chard
  • maritima are fertile and show compatibility at the chromosomal level (Forster et al., 1997). Hybrids between B. macrocarpa and B. vulgaris have caused weed problems in European sugar beet fields (McFarlane, 1975). Second, artificial hybrids have been produced with species in the section Corollinae. However, such hybrids are highly sterile and few plants set seed when backcrossed to sugar beet. Third, artificial hybrids between sugar beet and members of the genus Procumbentes have been produced with great difficulty. The hybrids become necrotic and die at the seedling stage. The chromosomes of the species of section Procumbentes do not pair with those of the genus Beta (Van Geyt et al., 1990).
  • the present disclosure teaches an interspecific cross of a beet of the present disclosure with another species in the genus Beta and/or an intergeneric cross of a beet of the present disclosure with another species not in the genus Be ta, but in the same family Amaranthaceae.
  • commercial lines of interest can be used as a receiver of desired trait(s) of the present disclosure by breeding and/or introgressing.
  • An exemplary list of the commercial lines are presented in Table 1.
  • particularly desirable traits that may be incorporated by this disclosure are improved resistance to different bacterial fungal, and viral pathogens.
  • Important diseases include but are not limited to (i) bacterial diseases such as Bacterial blight (Pseudomonas syringae pv. Apiaia). Bacterial pocket (Xanthomonas helicoid). Bacterial soft rot (Envinia carotovora subsp. Carotovora), Bacterial vascular necrosis and rot (Envinia carotovora subsp.
  • fungal disease such as Altemaria leaf spot (Alternaria alternata; Alternaria brassicae), Anthracnose (Colletotrichum dematium), Aphanomyces root rot (black root; Aphanomyces cochlioides).
  • Black wood vessel Pulcoal irregulare
  • Cercospora leaf spot Cercospora beticoki
  • Charcoal rot Macrophomina phaseolina
  • Choanephora rot Choanephora cucurbitarum
  • Damping-off black leg, black root and seedling blight (Aphanomyces cochlioides; Cylindrocladium spp.
  • Fusarium spp. Phoma betae Pleospora; betae; Pythium spp.; Rhizoctonia solani; Thanatephorus cucumeris), Downy mildew (Peronospora farinosa; Peronospora schachtii), Fusarium yellows (Fusarium oxysporum), Fusarium yellows and root rot (Fusarium oxysporum f.sp.
  • Insect pests affecting the various species of beet include Sugar beet root aphids (Pemphigus populivenae betae), Beet leaf miner (Pegomya hyoscyami), Beet webworms (Loxostege sticticalis), Blister beetles (Pyrota lineata), European Com Borers, Flea beetles (Psylliodes patheticata melsheimer), Sugar beet maggots (Tetanops myopaeformis) sugar beet nematodes (Heterodera schachiii)' and root not nematodes (Meloidogyne spp.) and vegetable weevils.
  • the present disclosure teaches additional desirable traits of disease resistance that can be combined with the yellowing vims resistant trait. That is, these additional desirable traits can be stacked with yellowing disease resistant traits taught herein.
  • the disease resistant traits include, but are not limited to, resistances against the following: Cercospora beticola Aphanomyces cochlioides, Heterodera schachtii, Beet necrotic yellow vein virus, Rhizoctonia solani, Pemphigus populivenae betae, Uromyces betae, Peronospora farinose, and Fusarium oxysporum.
  • biotechnological traits can be incorporated to the new beet plants of the present disclosure having resistance against yellowing vims(es).
  • the biotechnological traits include, but are not limited to, Roundup ready (H7-1), Acetolactate Synthase (ALS)- resistance, and Protoporphyrinogen Oxidase (PPO)-resistance.
  • CMS Cytosolic male sterility
  • GMS Genetic male sterility
  • Bolting resistance Hypocotyl colour
  • Genetic haploid induction systems extreme bolting tolerance (“winter beet”) and changes in carbon metabolism to achieve higher sugar contents.
  • the present disclosure teaches methods of using molecular markers taught herein for selection of BMYV -resistant plants.
  • the present disclosure teaches methods of selecting or identifying a plant comprising a resistance against Beet Mild Yellow Vims (BMYV), said method comprising the steps of: (a) screening for the presence of at least one QTL allele, such as a QTL allele associated with resistance against BMYV), wherein said QTL is selected from the group consisting of QTL1, QTL2, QTL3, and QTL4, wherein QTL1 located on chromosome 1 is genetically linked to at least one marker loci selected from the group consisting of PBV504245, PBV125552, PBV291217, PBV369685, PBV261963, PBV757437, PBV638722, and EPBV6296; wherein QTL2 located on chromosome 4 is genetically linked to at least one marker loci selected from the group consisting of PBV577030, PBV562771, PBV573138, PBV273480, and PBV821051, wherein QTL3 located on chromos
  • the screening for the presence of QTL1 comprises at least one of: PCR amplification of DNA with SEQ ID NO: 2 and SEQ ID NO: 3, PCR amplification of DNA with SEQ ID NO: 5 and SEQ ID NO: 6, PCR amplification of DNA with SEQ ID NO: 8 and SEQ ID NO: 9, PCR amplification of DNA with SEQ ID NO: 11 and SEQ ID NO: 12, PCR amplification of DNA with SEQ ID NO: 14 and SEQ ID NO: 15, PCR amplification of DNA with SEQ ID NO: 52 and SEQ ID NO: 54, PCR amplification of DNA with SEQ ID NO: 56 and SEQ ID NO: 57, and PCR amplification of DNA with SEQ ID NO: 58 and SEQ ID NO: 60.
  • the screening for the presence of QTL3 comprises at least one of: PCR amplification of DNA with SEQ ID NO: 32 and SEQ ID NO: 33, PCR amplification of DNA with SEQ ID NO: 35 and SEQ ID NO: 36, PCR amplification of DNA with SEQ ID NO: 38 and SEQ ID NO: 39, PCR amplification of DNA with SEQ ID NO: 41 and SEQ ID NO: 42, PCR amplification of DNA with SEQ ID NO: 44 and SEQ ID NO: 45, PCR amplification of DNA with SEQ ID NO: 62 and SEQ ID NO: 63, PCR amplification of DNA with SEQ ID NO: 65 and SEQ ID NO: 66, and PCR amplification of DNA with SEQ ID NO: 67 and SEQ ID NO: 69.
  • the screening for the presence of QTL4 comprises at least one of: PCR amplification of DNA with SEQ ID NO: 70 and SEQ ID NO: 72, PCR amplification of DNA with SEQ ID NO: 73 and SEQ ID NO: 75, PCR amplification of DNA with SEQ ID NO: 76 and SEQ ID NO: 78, PCR amplification of DNA with SEQ ID NO: 79 and SEQ ID NO: 81, PCR amplification of DNA with SEQ ID NO: 83 and SEQ ID NO: 84, and PCR amplification of DNA with SEQ ID NO: 85 and SEQ ID NO: 87.
  • a marker for selection of Beet Mild Yellowing Virus resistant plants is selected from the group consisting of SEQ ID NO: 1 to SEQ ID NO: 45 and SEQ ID NO: 52 to SEQ ID NO: 87.
  • the marker of the present disclosure utilizes a sequence selected from the group consisting of SEQ ID NO: 2, SEQ ID NO: 5, SEQ ID NO: 8, SEQ ID NO: 11, SEQ ID NO: 14, SEQ ID NO: 17, SEQ ID NO: 20, SEQ ID NO: 23, SEQ ID NO: 26, SEQ ID NO: 29, SEQ ID NO: 32, SEQ ID NO: 35, SEQ ID NO: 38, SEQ ID NO: 41, SEQ ID NO: 44, SEQ ID NO: 52, SEQ ID NO: 56, SEQ ID NO: 58, SEQ ID NO: 62, SEQ ID NO: 65, SEQ ID NO: 67, SEQ ID NO: 70, SEQ ID NO: 73, SEQ ID NO: 76, SEQ ID NO: 79, SEQ ID NO
  • the present disclosure teaches methods of producing a plant of the subspecies Beta vulgaris subsp. vulgaris expressing a resistance against Beet Mild Yellowing Virus (BMW), said method comprising the steps of: (a) providing a plant resistant to BMW, (b) crossing the plant of (a) with another plant and harvesting the seed resulting from said cross, (c) growing the seed from step (b). Also, methods comprise submitting the resulting plants for test, diagnosing, and selecting a plant which comprises at least one QTL selected from the group consisting of QTL1, QTL2, QTL3, and QTL4. Optionally, the method comprises (d) continuing the crossing until all QTLs are introgressed, and (e) further propagating or multiplying the resulting variety.
  • BMW Beet Mild Yellowing Virus
  • the present disclosure teaches methods for reducing yield loss as a consequence of BMW infection, the method comprising the steps of: (a) providing a plant resistant to BMW, a plant identified by the method taught herein, or a plant produced by the method taught herein, or a progeny of thereof having the resistance against BMW, (b) creating seed material, and (c) planting seed in areas which are prone to BMW infection.
  • the present disclosure teaches methods for producing sugar, the method comprising the steps of: (a) growing a plant according to the disclosure and/or a plant identified by the methods taught herein, or a plant produced by the method taught herein, or a progeny thereof having the having the resistance against BMYV, harvesting the root/beet of said plant, and (b) extracting the sugar from said root/beet of (b).
  • the growing in step (a) is conducted without the use of insecticides and the sugar extracted from said root/beet is qualified for the organic market.
  • the present disclosure teaches methods of identifying a nucleic acid molecule, which encodes a protein capable of imparting resistance to the pathogen BMYV in a plant of the Beta genus, in which the protein is expressed, characterized in that the method comprises the steps of: (a) detecting at least one polymorphism in the coding nucleotide sequence of the nucleic acid molecule taught herein, using molecular markers, which detect the polymorphism.
  • the present disclosure teaches that resistant varieties having at least one of QTLs taught herein may exhibit some disease symptoms or damage under heavy pest or pathogen pressure.
  • the viral load in the resistant variety is reduced by at least 5%, by at least 10%, by at least 15%, by at least 20%, by at least 25%, by at least 30%, by at least 35%, by at least 40%, by at least 45%, by at least 50%, by at least 100%, by at least 200%, by at least 30%, when compared to the susceptible variety or the wild type variety without at least one QTL taught herein.
  • the viral load is measured, for example, in an ELISA assay or qPCR.
  • the resistance against yellowing virus(es) is associated with a reduced load of the Beet Mild Yellows Virus by at least 25% in comparison to a variety lacking QTL1, QTL2, QTL3, and QTL4. In some embodiments, the resistance against yellowing virus(es) is associated with a reduced load of the Beet Mild Yellows Virus by at least 50% in comparison to a variety lacking QTL1, QTL2, QTL3, and QTL4. In some embodiments, the resistance against yellowing virus(es) is associated with a reduced load of the Beet Mild Yellows Virus by at least 75% in comparison to a variety lacking QTL1, QTL2, QTL3, and QTL4.
  • the present disclosure provides four QTLs on four different chromosomes; on chromosome 1, chromosome 4, chromosome 8, and chromosome 2, conferring tolerance or resistance to BMYV according to the present disclosure. These are heterozygously present in the genome of the plant deposited, and such plant can be used to produce additional BMYV resistant plants by self-crossing or out-crossing.
  • additional desirable traits may be introduced into the resistant or tolerant line. Methods of introducing additional desirable traits may include genetic transformation, genome editing, targeted mutagenesis, traditional breeding, and induced random mutagenesis.
  • Traits obtained by genetic transformations include - for example - (i) event H7-1 (OECD ID: KM-000H71-4), which confers resistance against glyphosate, and is described in US7335816, and is commercialized in for example the sugar beet varieties MA902 and HIL9528, (ii) event GTSB77 (OECD ID: SY-GTSB77-8) coding for glyphosate resistance in sugar beet, and (iii) event T120-7 (OECD ID: ACS-BV001-3) coding for glufosinate resistance in sugar beet, described in www.agbios.com/dbase.php# and www.isaaa.org/gmapprovaldatabase/default.asp.
  • Traits obtained by induced mutagenesis include - for example - tolerance against ALS inhibitor herbicide as described for example in EP3326453A1 (ALS INHIBITOR HERBICIDE TOLERANT BETA VULGARIS MUTANTS), US20210054360A1, and EP2627168A1, and available either from the related patent deposit (NCIMB, Aberdeen, UK, under Number NCIMB 41705) or from multiple commercially available sugar beet varieties including for example SMART RIXTA KWS.
  • EP3326453A1 ALS INHIBITOR HERBICIDE TOLERANT BETA VULGARIS MUTANTS
  • US20210054360A1 US20210054360A1
  • EP2627168A1 and available either from the related patent deposit (NCIMB, Aberdeen, UK, under Number NCIMB 41705) or from multiple commercially available sugar beet varieties including for example SMART RIXTA KWS.
  • the desirable trait is selected from the group consisting of (i) the H7-1 event conferring glyphosate tolerance, (ii) a mutated ALS gene conferring tolerance against ALS herbicides, (iii) the GTSB77 event conferring glyphosate tolerance, and (iv) the T120-7 event conferring glufosinate tolerance.
  • the ALS gene encodes an ALS polypeptide containing an amino acid different from tryptophan at a position 569 of the ALS polypeptide.
  • a deposit of the sugar beet seed of this disclosure is maintained by DLF Seeds A/S (Ny Oestergade 9, Roskilde 4000, Denmark).
  • a sample of the beet seed of this disclosure has been deposited with the National Collections of Industrial, Food and Marine Bacteria (NCIMB), 23 St Machar Drive, Aberdeen, Scotland, AB24 3RY, United Kingdom.
  • NCIMB National Collections of Industrial, Food and Marine Bacteria
  • the seed deposit of the beet cultivar 21000003-71 was made on January 27, 2023.
  • the QTL associated with tolerance to BMYV described herein are chosen from those present in the genome of sugar beet line 21000003-71, which seeds are deposited under the NCIMB accession number 44107.
  • the Beta vulgaris subsp. vulgaris plant according to the disclosure is line 21000003-71, which seeds are deposited under NCIMB accession number 44107.
  • the deposited beet cultivar 21000003-71 is a uniform Fl hybrid, which is heterozygous for all four QTLs (i.e., QTL1, QTL2, QTL3, and QTL4 taught herein).
  • Beet cultivar 21000003-71 was developed by the introgression of a donor beet variety having four QTLs associated with resistance to BMYV into a susceptible beet variety or a beet variety lacking the QTLs taught herein.
  • Beet cultivar 21000003-71 can be used as breeding material and also commercialized as organic commodity plant products.
  • BC2 back-cross population 2
  • the identified genetic determinants were obtained from a private collection of wild beet, which is proprietarily owned by DLF Seeds A/S and not publicly available. A total of 200 individuals from this BC2 was analyzed using 15 trait specific markers. 10 plants that were heterozygous for the traits were selected and self-crossed to create a BC2S 1 -population that were segregating for all three genetic determinants.
  • a total of 1200 plants were analyzed for the presence of the genetic determinants and one or more plants that were homozygous for the trait were selected. Further self-crossing was conducted until an acceptable level of line homozygosity was achieved and the resulting line is regarded as an output from the introgression program. From each initial cross between the resistant and susceptible germplasm, several outputs were produced and evaluated for resistance.
  • the aphids were killed with an insecticide four days after inoculation and the plants were sprayed with insecticides several times over the growing season to avoid contamination by other aphids.
  • the development of symptoms was scored every 3 rd week during the entire season.
  • the highest concentration in the standard curve is 0.15 g freeze dried powder mixed with 3 ml extraction buffer (“100%” in the standard curve ).
  • the standard curve was made in 10 steps and for each step the sample was diluted with buffer 1 : 1 giving the concentrations: 100%, 50%, 25%, 12.5% etc.
  • DSMZ Leibniz Institute DSMZ -German Collection of Microorganisms and Cell Cultures GmbH
  • the antisera for BMYV and BChV is unable to distinguish between the two poleroviruses BMYV and BChV.
  • RNAlater stabilization solution ThermoFischer Scientific
  • RNAlater stabilization solution ThermoFischer Scientific
  • concentration of the RNA was then measured and all samples were diluted to 80 ng/pl before cDNA synthesis was carried out using the iScript cDNA Synthesis kit (BIORAD) following the manufacturer’s instructions.
  • qRT-PCR was then set up and ran using the following program: 95°C for 10 min followed by 95°C for 15 seconds, 54°C for 15 seconds, 72°C for 20 seconds for 40 cycles. Three pairs of virus specific primers were used (Table 2).
  • Table 3 shows the result of BMYV-infection in semi -field trial conditions on three different elite germplasm before and after the introgression of the resistance trait described above.
  • LineX (Res) denotes the introgressed version of “LineX (Sus)” into which all four genetic determinants have been introgressed and fixed in a homozygous form.
  • Control is a susceptible line. This shows that it is possible to transfer the resistance trait using the methods described herein and that the resistance is largely independent of the receiving germplasm, although some minor variation is observable which is to be expected.
  • Table 4 shows ELISA %BMYV data for leaves of different ages (“Old”, “Middle” and “Young”) sampled at two different timepoints (TO and 21 days after TO).
  • “Source” is the original resistance source used for the identification of the genetic determinants.
  • “LineX (Res)” denotes the introgressed version of “LineX (Sus)” into which all four genetic determinants have been introgressed using the method described above.
  • Line B the numbers denote different outputs from the same introgression project. This shows that the introgressed resistance trait is systemic in the plant and stable over time and not significantly influenced by sampling time point. Table 4
  • Three-way hybrids can be created in a number of ways but most commonly a male sterility system is used allowing for large scale production of tightly controlled crosses.
  • the first step is to convert a fertile elite germplasm into a male sterile by transferring the genetic determinants required for the male sterility trait.
  • This male sterile germplasm can then be pollinated by a fully fertile line creating a heterozygous but heterogenous Fl population. If for example a cytosolic male sterility system is used, the male sterility can also be maintained in the Fl. This Fl MS can then easily be pollinated by an additional pollinator to create the final three-way hybrid.
  • Dominant traits can be introduced and maintained in either the female (F 1 MS) side or the male (pollinator) side of the final hybrid while a recessive trait must be fixed in all three components of the final hybrid.
  • a dominant trait is therefore of very high economic value to the breeders.
  • the genetic determinants were introgressed into several pollinator germplasm and combined with several commercially relevant F1MS- mothers. These three-way hybrids were then evaluated in semi-field and field conditions for virus yellows resistance.
  • Tables 5 and 6 contains data from semi-field evaluation of three different three-way hybrids showing the results from phenotypic scoring and ELISA-based quantification of virus content for all three viruses.
  • Table 5 covers experiments carried out in year 1 (2021) while Table 6 contains data from year 3 (2023).
  • “Pollinator (Res)” is the resistant pollinator created by introgressing the genetic determinants from “Source” into “Control (Pollinator (Sus))”.
  • F1MS-X is the single hybrid used as the parent for the three-way hybrid. This shows that the resistance achievable by the above described method of introgression is functional in a hybrid and heterozygous form against BMYV and BChV. No data was collected for BYV except for the introgressed pollinator (line level) showing that some resistance/tolerance against BYV is linked to the same genetic determinants.
  • ND No data.
  • the resistant hybrids (#1, #3) show a clear improvement in phenotype (Score) and a substantial reduction in virus content in comparison to non-resistant/non-tolerant material (#2) but also in comparison to varieties Maruscha (#5) and Yellowstone (#4) which are described to be BMYV-tolerant.
  • Table 10 contains results from yield trials conducted in Svalof, Sweden in year 2 using a 20% infection rate. All entries, with the exceptions from the two controls (Y ellowstone and Maruscha) which are commercially available hybrids, consists of three-way hybrids consisting of the same female (F1MS-D) crossed with a pollinator.
  • the pollinator group DI -D6 represents closely related germplasm where an elite pollinator (DI) with no resistance against BMYV and BYV has undergone the introgression process to transfer the genetic determinants with the resulting outputs (D2-D6) used for hybrid production.
  • Pollinator El and E2 represent a similar setup but with only a single output (E2). The % yield was calculated using the following formula:
  • the resistant hybrids (#1, #3, #4, #5, #6, #7) show an improvement in yield in comparison to non-resistant/non-tolerant material (#2, #8, #9) under the conditions of a moderate (20%) infection challenge.
  • Table 12 shows data from official trials in Belgium Year 2 using a 5% infection rate where field inoculated with BMYV were scored at three different timepoints: 32, 43 and 71 days post inoculation (DPI). Scoring was done by counting number of plants with visible symptoms and them calculating their percentage. The same single hybrid was used as the mother for all three hybrids. This result confirms that the resistant hybrid carrying the trait in a heterozygous form is able to withstand the disease pressure significantly better than the susceptible hybrids.
  • DPI days post inoculation
  • Table 13 contains data from UK COMEX trials in 2023 where a 100% infection rate of all three viruses (BYC, BMYV and BChV) was used and the yield reduction measured.
  • the resistant hybrids (#3, #4, #5, #6) show an improvement in yield in comparison to non-resistant/non-tolerant material (#1, #2).
  • a homozygous resistant line (“donor parent”) carrying the four required genetic determinants is crossed with a susceptible line (’’recurrent parent”).
  • a recurrent parent can consist of any susceptible individual belonging to a species crossable with sugar beet such as members of the Beta vulgaris ssp. vulgaris group which includes crops such as sugar beet, chard, beetroot and mangold but also closely related species such as Beta vulgaris ssp. maritima or any other member of the Betoideae family able to produce viable offspring when crossed with Beta vulgaris ssp. vulgaris.
  • the resulting Fl can then be self-crossed to establish an F2 population in which the genetic determinants are segregating.
  • individuals that carry one or more of these genetic determinants This process can then be repeated with additional crosses to resistant germplasm until all three genetic determinants have been assembled and the resistant trait has been introgressed into the selected germplasm.
  • Another approach is to backcross the Fl with the recurrent parent. Single plant offspring from this first backcross (BC1) or any additional backcrosses (BCX) can then be selected using the markers developed from the initial mapping population to identify plants carrying the genetic determinants responsible for the trait in a heterozygous or homozygous form.
  • plants carrying the markers for the trait but otherwise showing high similarity with the receiver line are selected for selfing. These plants can then be selfcrossed to produce a BCXS 1 which will be segregating for the desired trait.
  • plants which carry the trait in either heterozygous or homozygous form can be selected for further breeding and evaluation.
  • a large number of the offspring is then analyzed using the molecular markers described above to determine the inheritance pattern across the genome between the donor and receiver parental genotypes. In this analysis, a number (-100 SNP-markers) are used to trace the parental genomes over a population of around 100 offsprings and identify the individuals carrying the desired regions associated with the traits from the donor parent.
  • Flanking markers for the trait have been developed to allow for the identification of crossovers close to the region of interest. Additional markers (i.e. five) between the two flanking markers are also used to ensure correct introgression. Fewer markers can be used but increase the risk of failed introgression due to unwanted crossovers. Other markers are used to select for crossovers with the receiver germplasm. Based on these markers, plants carrying the markers for the trait but otherwise showing high similarity with the receiver line are selected for selfing. At this stage, only heterozygous trait specific markers are selected.
  • the resulting heterozygous offspring can be used to create a segregating population by additional crossing using any suitable germplasm, including self-crossing.
  • any suitable germplasm including self-crossing.
  • homozygous resistant germplasm can be extracted by using the described molecular markers. For this, plants from the segregating population are analyzed for the presence of the resistant alleles and the plants carrying all the correct alleles are selected as resistant.
  • the fixation of all genetic determinants can be done stepwise by repeated crosses. For the purpose of marker selection, both the confidence interval (usually 95%) or the single point best likelihood model can be utilized.
  • KASP genotyping is based on competitive allele-specific PCR and is within this application used to distinguish germplasm carrying the relevant genetic determinants in a bi- allelic manner.
  • the KASP-technology requires two allele-specific forward primers and one common reverse primer.
  • Each allele-specific primer carries a FRET (Forster resonance energy transfer) cassette labeled with either the FAM or HEX dye.
  • FRET cassettes are associated through sequence homology with a quencher molecule. Once the newly synthesized template is available, the quencher carrying sequence will dissociate allowing the FRET cassette allowing the fluorophores (HEX and FAM) to emit light within their respective emission range.
  • FRET Forward resonance energy transfer
  • SEQ ID NO: 1-45, and SEQ ID NO: 52 to SEQ ID NO: 87 are primer sequences used for KASP/marker analysis of the trait.
  • Three primers are needed for the marker system to work (e.g., SEQ ID NO: 1-3).
  • SEQ ID NO: 1 is the forward primer for the susceptible allele at that position
  • SEQ ID NO:2 is the forward primer for the resistant allele
  • SEQ ID NO:3 is the common reverse primer for both forward primers.
  • all three primers would be mixed, but theoretically, only the primer for the resistant allele is required to detect the SNP of interest. If only one primer is used, heterozygous alleles are not able to be detected (the assay becomes dominant).
  • SEQ ID NO:46-SEQ ID NO:51 are primers used in qRT-PCR verification ofthe viruses and are also presented in Table 2.
  • SEQ ID NO:46 and SEQ ID NO:47 are specific for BMYV
  • SEQ ID NO:48 and SEQ ID NO:49 are specific for BChV
  • SEQ ID NO:50 and SEQ ID NO:51 are specific for BYV.
  • the PCR condition is described in Table 15 and the primer sequence information in Table 16.
  • Example 9 - Resistant lines disclosed herein exhibit significantly reduced viral load compared to lines lacking OTLl, QTL2, QTL3, and QTL4
  • the varieties provided by KWS provide a certain tolerance against yellowing viruses, which mitigates some of the effects to some extent and provides a certain “stay green” effect under virus pressure.
  • no substantial reduction of virus load can be seen from Marushca KWS when compared to a hybrid of the present disclosure containing three QTLs (i.e., QTL1, QTL2, QTL3, and QTL4), as presented in Table 17.
  • the value shown in Table 17 is the average ELISA-results of 8 plants from semi-field trials to test resistance to yellowing viruses (BMYV, BYV, and BChV).
  • the susceptible hybrid plants as a control show no resistance to yellowing viruses and a full viral load.
  • Maruscha KWS plants show mild tolerance to yellowing viruses with no significant reduction of viral load (about 20%)
  • the resistant hybrid plants possessing three QTLs of the present disclosure present strong resistance with significant reduction of viral road (about 66% to about 99%), which is at least about 3 times higher resistance than the Maruscha KWS.
  • Beta vulgaris subsp. vulgaris plant, plant part, or plant cell resistant to Beet Mild Yellowing Virus (BMYV) 1.
  • QTL Quantitative Trait Loci
  • SNP single nucleotide polymorphism
  • PBV504245 is a C to G substitution corresponding to position 1050427 of chromosome
  • PBV125552 is a G to A substitution corresponding to position 3069972 of chromosome
  • PBV291217 is a G to A substitution corresponding to position 4315236 of chromosome
  • PBV369685 is an A to G substitution corresponding to position 4330737 of chromosome 1 of the Refbeet 1.5 reference genome
  • PBV261963 is an A to T substitution corresponding to position 4356001 of chromosome 1 of the Refbeet 1.5 reference genome
  • SNP single nucleotide polymorphism
  • PBV757437 is a G to A substitution corresponding to position 17043 of chromosome 1 of the Refbeet 1.5 reference genome
  • PBV638722 is an A to G substitution corresponding to position 2645728 of chromosome
  • EPBV6296 is a T to C substitution corresponding to position 1117040 of chromosome 1 of the Refbeet 1.5 reference genome.
  • SNP single nucleotide polymorphism
  • PBV562771 is a G to A substitution corresponding to position 9767085 of chromosome
  • PBV573138 is an A to G substitution corresponding to position 10271308 of chromosome 4 of the Refbeet 1.5 reference genome
  • PBV273480 is a G to A substitution corresponding to position 46718199 of chromosome 4 of the Refbeet 1.5 reference genome
  • PBV821051 is a G to A substitution corresponding to position 47855576 of chromosome 4 of the Refbeet 1.5 reference genome.
  • SNP single nucleotide polymorphism
  • PBV148969 is an A to G substitution corresponding to position 436005 of chromosome
  • PBV331644 is a G to A substitution corresponding to position 1366650 of chromosome
  • PBV882826 is an A to G substitution corresponding to position 7979633 of chromosome 8 of the Refbeet 1.5 reference genome
  • PBV374371 is a G to A substitution corresponding to position 8822544 of chromosome
  • PBV757436 is an A to C substitution corresponding to position 25201555 of chromosome 8 of the Refbeet 1.5 reference genome.
  • SNP single nucleotide polymorphism
  • PBV738292 is a G to an A substitution corresponding to position 35831 of chromosome
  • PBV738293 is an A to G substitution corresponding to position 1366650 of chromosome 8 of the Refbeet 1.5 reference genome
  • EPBV7349 is an A to C substitution corresponding to position 275340 of chromosome
  • GRKPBV98281 is a G to A substitution corresponding to position 8040503 of chromosome 2 of the Refbeet 1.5 reference genome
  • GRKPBV627372 is a C to an A substitution corresponding to position 9433027 of chromosome 2 of the Refbeet 1.5 reference genome
  • GRKPBV672273 is a G to an A substitution corresponding to position 12212298 of chromosome 2 of the Refbeet 1.5 reference genome
  • GRKPBV627178 is an A to C substitution corresponding to position 18410288 of chromosome 2 of the Refbeet 1.5 reference genome
  • GRKPBV116271 is an A to C substitution corresponding to position 20007280 of chromosome 2 of the Refbeet 1.5 reference genome, and wherein QTL4 is obtainable from Beta vulgaris subsp. vulgaris line 21000003-71, representative seed of which has been deposited under NCIMB number 44107.
  • ALS gene encodes an ALS polypeptide containing an amino acid different from tryptophan at a position 569 of the ALS polypeptide.
  • a marker for selection of Beet Mild Yellowing Virus resistant plants selected from the group consisting of SEQ ID NO: 1 to SEQ ID NO: 45.
  • a marker for selection of Beet Mild Yellowing Virus resistant plants selected from the group consisting of SEQ ID NO: 52 to SEQ ID NO: 87.
  • the marker of embodiment 49 wherein the marker utilizes a sequence selected from the group consisting of SEQ ID NO: 2, SEQ ID NO: 5, SEQ ID NO: 8, SEQ ID NO: 11, SEQ ID NO: 14, SEQ ID NO: 17, SEQ ID NO: 20, SEQ ID NO: 23, SEQ ID NO: 26, SEQ ID NO: 29, SEQ ID NO: 32, SEQ ID NO: 35, SEQ ID NO: 38, SEQ ID NO: 41, and SEQ ID NO: 44.
  • the marker of embodiment 50 wherein the marker utilizes a sequence selected from the group consisting of SEQ ID NO: 52, SEQ ID NO: 56, SEQ ID NO: 58, SEQ ID NO: 62, SEQ ID NO: 65, SEQ ID NO: 67, SEQ ID NO: 70, SEQ ID NO: 73, SEQ ID NO: 76, SEQ ID NO: 79, SEQ ID NO: 83, and SEQ ID NO: 85.
  • BMYV Beet Mild Yellowing Virus
  • BYV Beet yellows virus
  • BChV Beet chlorosis virus
  • molecular markers identify at least one of the Single Nucleotide Polymorphisms present in PBV757437, PBV638722, EPBV6296, PBV738292, PBV738293, EPBV7349, GRKPBV98281, GRKPBV627372, GRKPBV672273, GRKPBV627178, GRKPBV116271, and EPBV8821.
  • a method of identifying a plant, plant part, or plant cell comprising a Quantitative Trait Locus (QTL) associated with resistance to Beet Mild Yellow Virus (BMYV) on chromosome 1 (QTL1), said method comprising the steps of: screening for the presence of QTL1, wherein QTL1 is genetically linked to at least one single nucleotide polymorphism (SNP) selected from the group consisting of PBV504245, PBV125552, PBV291217, PBV369685, and PBV261963, wherein:
  • PBV504245 is a C to G substitution corresponding to position 1050427 of chromosome 1 of the Refbeet 1.5 reference genome
  • PBV125552 is a G to A substitution corresponding to position 3069972 of chromosome 1 of the Refbeet 1.5 reference genome
  • PBV291217 is a G to A substitution corresponding to position 4315236 of chromosome 1 of the Refbeet 1.5 reference genome
  • PBV369685 is an A to G substitution corresponding to position 4330737 of chromosome 1 of the Refbeet 1.5 reference genome
  • PBV261963 is an A to T substitution corresponding to position 4356001 of chromosome 1 of the Refbeet 1.5 reference genome, and identifying a plant, plant part, or plant cell having at least one of said marker loci linked to QTL1.
  • said screening for the presence of QTL1 further comprises identifying a plant, plant part, or plant cell having at least one single nucleotide polymorphism (SNP) selected from the group consisting of PBV757437, PBV638722, and EPBV6296, wherein:
  • SNP single nucleotide polymorphism
  • PBV757437 is a G to A substitution corresponding to position 17043 of chromosome 1 of the Refbeet 1.5 reference genome
  • PBV638722 is an A to G substitution corresponding to position 2645728 of chromosome 1 of the Refbeet 1.5 reference genome
  • EPBV6296 is a T to C substitution corresponding to position 1117040 of chromosome 1 of the Refbeet 1.5 reference genome.
  • a method of identifying a plant, plant part, or plant cell comprising a Quantitative Trait Locus (QTL) associated with resistance to Beet Mild Yellow Virus (BMYV) on chromosome 4 (QTL2), said method comprising the steps of: screening for the presence of QTL2, wherein QTL2 is genetically linked to at least one single nucleotide polymorphism (SNP) selected from the group consisting of PBV577030, PBV562771, PBV573138, PBV273480, and PBV821051, wherein:
  • PBV577030 is an A to G substitution corresponding to position 5742753 of chromosome 4 of the Refbeet 1.5 reference genome
  • PBV562771 is a G to A substitution corresponding to position 9767085 of chromosome 4 of the Refbeet 1.5 reference genome
  • PBV573138 is an A to G substitution corresponding to position 10271308 of chromosome 4 of the Refbeet 1.5 reference genome
  • PBV273480 is a G to A substitution corresponding to position 46718199 of chromosome 4 of the Refbeet 1.5 reference genome
  • PBV821051 is a G to A substitution corresponding to position 47855576 of chromosome 4 of the Refbeet 1.5 reference genome, and identifying a plant, plant part, or plant cell having at least one of said marker loci linked to QTL2.
  • a method of identifying a plant, plant part, or plant cell comprising a Quantitative Trait Locus (QTL) associated with resistance to Beet Mild Yellow Virus (BMYV) on chromosome 8 (QTL3), said method comprising the steps of: screening for the presence of QTL3, wherein QTL3 is genetically linked to at least one single nucleotide polymorphism (SNP) selected from the group consisting of PBV148969, PBV331644, PBV882826, PBV374371, and PBV757436, wherein: PBV148969 is an A to G substitution corresponding to position 436005 of chromosome 8 of the Refbeet 1.5 reference genome,
  • QTL3 is genetically linked to at least one single nucleotide polymorphism (SNP) selected from the group consisting of PBV148969, PBV331644, PBV882826, PBV374371, and PBV757436, wherein: PBV148969 is an A to G substitution
  • PBV331644 is a G to A substitution corresponding to position 1366650 of chromosome 8 of the Refbeet 1.5 reference genome
  • PBV882826 is an A to G substitution corresponding to position 7979633 of chromosome 8 of the Refbeet 1.5 reference genome
  • PBV374371 is a G to A substitution corresponding to position 8822544 of chromosome 8 of the Refbeet 1.5 reference genome
  • PBV757436 is an A to C substitution corresponding to position 25201555 of chromosome 8 of the Refbeet 1.5 reference genome, and identifying a plant, plant part, or plant cell having at least one of said marker loci linked to QTL3.
  • PBV738292 is a G to an A substitution corresponding to position 35831 of chromosome 8 of the Refbeet 1.5 reference genome
  • PBV738293 is an A to G substitution corresponding to position 1366650 of chromosome 8 of the Refbeet 1.5 reference genome
  • EPBV7349 is an A to C substitution corresponding to position 275340 of chromosome 8 of the Refbeet 1.5 reference genome.
  • a method of identifying a plant, plant part, or plant cell comprising a Quantitative Trait Locus (QTL) associated with resistance to Beet Mild Yellow Virus (BMYV) on chromosome 2 (QTL4), said method comprising the steps of: screening for the presence of QTL4, wherein QTL4 is genetically linked to at least one single nucleotide polymorphism (SNP) selected from the group consisting of GRKPBV98281, GRKPBV627372, GRKPBV672273, GRKPBV627178, GRKPBV116271, and EPBV8821, wherein:
  • GRKPBV98281 is a G to A substitution corresponding to position 8040503 of chromosome 2 of the Refbeet 1.5 reference genome
  • GRKPBV627372 is a C to an A substitution corresponding to position 9433027 of chromosome 2 of the Refbeet 1.5 reference genome
  • GRKPBV672273 is a G to an A substitution corresponding to position 12212298 of chromosome 2 of the Refbeet 1.5 reference genome
  • GRKPBV627178 is an Ato C substitution corresponding to position 18410288 of chromosome 2 of the Refbeet 1.5 reference genome
  • GRKPBV 116271 is an Ato C substitution corresponding to position 20007280 of chromosome 2 of the Refbeet 1.5 reference genome, and identifying a plant, plant part, or plant cell having at least one of said marker loci linked to QTL4.
  • BMYV Beet Mild Yellowing Virus
  • step (c) growing the seed from step (b) to produce a progeny plant, and submitting the progeny plant to the method of identifying a plant comprising at least one of a Quantitative Trait Locus (QTL) associated with resistance to Beet Mild Yellow Virus (BMYV); and
  • QTL Quantitative Trait Locus
  • selecting further comprises selecting a progeny plant having QTL4.
  • PCR amplification of DNA with SEQ ID NO: 5 and SEQ ID NO: 6 PCR amplification of DNA with SEQ ID NO: 8 and SEQ ID NO: 9, PCR amplification of DNA with SEQ ID NO: 11 and SEQ ID NO: 12, and PCR amplification of DNA with SEQ ID NO: 14 and SEQ ID NO: 15.
  • PCR amplification of DNA with SEQ ID NO: 17 and SEQ ID NO: 18 PCR amplification of DNA with SEQ ID NO: 20 and SEQ ID NO: 21, PCR amplification of DNA with SEQ ID NO: 23 and SEQ ID NO: 24, PCR amplification of DNA with SEQ ID NO: 26 and SEQ ID NO: 27, and PCR amplification of DNA with SEQ ID NO: 29 and SEQ ID NO: 30.
  • PCR amplification of DNA with SEQ ID NO: 32 and SEQ ID NO: 33 PCR amplification of DNA with SEQ ID NO: 35 and SEQ ID NO: 36, PCR amplification of DNA with SEQ ID NO: 38 and SEQ ID NO: 39, PCR amplification of DNA with SEQ ID NO: 41 and SEQ ID NO: 42, and PCR amplification of DNA with SEQ ID NO: 44 and SEQ ID NO: 45.
  • PCR amplification of DNA with SEQ ID NO: 70 and SEQ ID NO: 72 PCR amplification of DNA with SEQ ID NO: 73 and SEQ ID NO: 75
  • PCR amplification of DNA with SEQ ID NO: 76 and SEQ ID NO: 78 PCR amplification of DNA with SEQ ID NO: 79 and SEQ ID NO: 81
  • PCR amplification of DNA with SEQ ID NO: 83 and SEQ ID NO: 84 and PCR amplification of DNA with SEQ ID NO: 85 and SEQ ID NO: 87.
  • invention 80 The method of embodiment 78 or 79, wherein said desirable trait is selected from the group consisting of (i) the H7-1 event conferring glyphosate tolerance, (ii) a mutated Acetolactate Synthase (ALS) gene conferring tolerance against ALS herbicides, (iii) the GTSB77 event conferring glyphosate tolerance, and (iv) the T120-7 event conferring glufosinate tolerance.
  • ALS Acetolactate Synthase
  • a method for reducing yield loss as a consequence of Beet Mild Yellowing Virus (BMYV) infection comprising the steps of:
  • a method for producing sugar comprising the steps of:

Landscapes

  • Health & Medical Sciences (AREA)
  • Life Sciences & Earth Sciences (AREA)
  • Botany (AREA)
  • Genetics & Genomics (AREA)
  • Developmental Biology & Embryology (AREA)
  • Environmental Sciences (AREA)
  • General Health & Medical Sciences (AREA)
  • Spectroscopy & Molecular Physics (AREA)
  • Physics & Mathematics (AREA)
  • Natural Medicines & Medicinal Plants (AREA)
  • Physiology (AREA)
  • Breeding Of Plants And Reproduction By Means Of Culturing (AREA)
  • Micro-Organisms Or Cultivation Processes Thereof (AREA)
  • Measuring Or Testing Involving Enzymes Or Micro-Organisms (AREA)

Abstract

La présente invention concerne des plantes, des parties de plante ou des cellules de plante résistantes au virus de la jaunisse modéré de la betterave (BMYV - Beet Mild Yellowing Virus) ayant au moins un locus de caractère quantitatif (QTL) associé à la résistance au BMYV. L'invention concerne également des procédés de sélection, d'identification et de production de plantes, de parties de plante ou de résistance de cellules végétales au BMYV, qui réduisent la perte de rendement ou augmentent le rendement de production de sucre. En outre, la présente invention concerne des molécules d'acide nucléique qui confèrent une résistance à un pathogène dans des plantes, des parties de plante et des cellules végétales, en particulier du genre Beta.
PCT/IB2024/050952 2023-02-01 2024-02-01 Résistance au virus de la jaunisse des betteraves Ceased WO2024161358A1 (fr)

Priority Applications (1)

Application Number Priority Date Filing Date Title
EP24703866.4A EP4658061A1 (fr) 2023-02-01 2024-02-01 Résistance au virus de la jaunisse des betteraves

Applications Claiming Priority (2)

Application Number Priority Date Filing Date Title
US202363482786P 2023-02-01 2023-02-01
US63/482,786 2023-02-01

Publications (1)

Publication Number Publication Date
WO2024161358A1 true WO2024161358A1 (fr) 2024-08-08

Family

ID=89853539

Family Applications (1)

Application Number Title Priority Date Filing Date
PCT/IB2024/050952 Ceased WO2024161358A1 (fr) 2023-02-01 2024-02-01 Résistance au virus de la jaunisse des betteraves

Country Status (2)

Country Link
EP (1) EP4658061A1 (fr)
WO (1) WO2024161358A1 (fr)

Cited By (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
JP7724484B1 (ja) * 2024-03-11 2025-08-18 国立研究開発法人農業・食品産業技術総合研究機構 テンサイ植物の萎黄性ウイルス病抵抗性マーカー及びその利用

Citations (14)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US5994075A (en) 1996-05-17 1999-11-30 Hexagen Technology Limited Methods for identifying a mutation in a gene of interest without a phenotypic guide
US20040053236A1 (en) 2001-03-30 2004-03-18 Mccallum Claire M. Reverse genetic strategy for identifying functional mutations in genes of known sequences
WO2005048692A2 (fr) 2003-11-17 2005-06-02 Arcadia Biosciences, Inc. Tomates a activite desoxyhypusine synthase reduite par suite d'alterations non transgeniques du gene de la desoxyhypusine synthase
WO2005055704A2 (fr) 2003-12-03 2005-06-23 Arcadia Biosciences, Inc. Ble comprenant une proteine cireuse reduite du fait d'alterations non transgeniques d'un gene cireux
US7335816B2 (en) 2003-02-28 2008-02-26 Kws Saat Ag Glyphosate tolerant sugar beet
US8440432B2 (en) 2009-12-10 2013-05-14 Regents Of The University Of Minnesota Tal effector-mediated DNA modification
EP2627168A1 (fr) 2010-10-15 2013-08-21 Bayer Intellectual Property GmbH Mutants beta vulgaris tolérants à un herbicide inhibiteur d'als
US8586526B2 (en) 2010-05-17 2013-11-19 Sangamo Biosciences, Inc. DNA-binding proteins and uses thereof
US8592645B2 (en) 2009-10-22 2013-11-26 Dow Agrosciences Llc Engineered zinc finger proteins targeting plant genes involved in fatty acid biosynthesis
US8697359B1 (en) 2012-12-12 2014-04-15 The Broad Institute, Inc. CRISPR-Cas systems and methods for altering expression of gene products
US8704041B2 (en) 2009-12-30 2014-04-22 Pioneer Hi Bred International Inc Methods and compositions for targeted polynucleotide modification
US8871445B2 (en) 2012-12-12 2014-10-28 The Broad Institute Inc. CRISPR-Cas component systems, methods and compositions for sequence manipulation
US8889418B2 (en) 2012-12-12 2014-11-18 The Broad Institute Inc. Engineering and optimization of improved systems, methods and enzyme compositions for sequence manipulation
US8921112B2 (en) 2006-12-14 2014-12-30 Dow Agrosciences Llc Optimized non-canonical zinc finger proteins

Patent Citations (21)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US5994075A (en) 1996-05-17 1999-11-30 Hexagen Technology Limited Methods for identifying a mutation in a gene of interest without a phenotypic guide
US20040053236A1 (en) 2001-03-30 2004-03-18 Mccallum Claire M. Reverse genetic strategy for identifying functional mutations in genes of known sequences
US7335816B2 (en) 2003-02-28 2008-02-26 Kws Saat Ag Glyphosate tolerant sugar beet
WO2005048692A2 (fr) 2003-11-17 2005-06-02 Arcadia Biosciences, Inc. Tomates a activite desoxyhypusine synthase reduite par suite d'alterations non transgeniques du gene de la desoxyhypusine synthase
WO2005055704A2 (fr) 2003-12-03 2005-06-23 Arcadia Biosciences, Inc. Ble comprenant une proteine cireuse reduite du fait d'alterations non transgeniques d'un gene cireux
US8921112B2 (en) 2006-12-14 2014-12-30 Dow Agrosciences Llc Optimized non-canonical zinc finger proteins
US8592645B2 (en) 2009-10-22 2013-11-26 Dow Agrosciences Llc Engineered zinc finger proteins targeting plant genes involved in fatty acid biosynthesis
US8440432B2 (en) 2009-12-10 2013-05-14 Regents Of The University Of Minnesota Tal effector-mediated DNA modification
US8586363B2 (en) 2009-12-10 2013-11-19 Regents Of The University Of Minnesota TAL effector-mediated DNA modification
US8450471B2 (en) 2009-12-10 2013-05-28 Regents Of The University Of Minnesota TAL effector-mediated DNA modification
US8697853B2 (en) 2009-12-10 2014-04-15 Regents Of The University Of Minnesota TAL effector-mediated DNA modification
US8704041B2 (en) 2009-12-30 2014-04-22 Pioneer Hi Bred International Inc Methods and compositions for targeted polynucleotide modification
US8912138B2 (en) 2010-05-17 2014-12-16 Sangamo Biosciences, Inc. DNA-binding proteins and uses thereof
US8586526B2 (en) 2010-05-17 2013-11-19 Sangamo Biosciences, Inc. DNA-binding proteins and uses thereof
EP2627168A1 (fr) 2010-10-15 2013-08-21 Bayer Intellectual Property GmbH Mutants beta vulgaris tolérants à un herbicide inhibiteur d'als
EP3326453A1 (fr) 2010-10-15 2018-05-30 Bayer Intellectual Property GmbH Mutants bêta vulgaris tolérants à l'herbicide inhibiteur als
US20210054360A1 (en) 2010-10-15 2021-02-25 Bayer Intellectual Property Gmbh Als inhibitor herbicide tolerant beta vulgaris mutants
US8697359B1 (en) 2012-12-12 2014-04-15 The Broad Institute, Inc. CRISPR-Cas systems and methods for altering expression of gene products
US8771945B1 (en) 2012-12-12 2014-07-08 The Broad Institute, Inc. CRISPR-Cas systems and methods for altering expression of gene products
US8871445B2 (en) 2012-12-12 2014-10-28 The Broad Institute Inc. CRISPR-Cas component systems, methods and compositions for sequence manipulation
US8889418B2 (en) 2012-12-12 2014-11-18 The Broad Institute Inc. Engineering and optimization of improved systems, methods and enzyme compositions for sequence manipulation

Non-Patent Citations (56)

* Cited by examiner, † Cited by third party
Title
ALLARD: "Principles of Plant Breeding", 1960, JOHN WILEY & SONS, INC.
ALLEN ET AL., PLANT BIOTECHNOLOGY J., vol. 9, 2011, pages 1086 - 1099
ALTSCHUL ET AL., J. MOL. BIOL., vol. 215, 1990, pages 403 - 10
ANNIKA HJERDIN PANAGOPOULOS, APPLICATION OF MOLECULAR MARKERS IN SUGAR BEET BREEDING, 2003, Retrieved from the Internet <URL:lucris.lub.lu.se/ws/portalfiles/portal/5776296/1693169.pdf>
BENNETT ET AL.: "Effects of virus yellows on sugar beet with a consideration of some of the factors involved in changes produced by the disease", AMERICAN SOCIETY OF SUGAR BEET TECHNOLOGISTS JOURNAL, vol. 9, 1956, pages 479 - 494
COMAI ET AL., THE PLANT JOURNAL, vol. 37, 2003, pages 778 - 786
DE KOEIJERVAN DER WERF: "Effects of Beet yellows virus and Beet mild yellowing virus on leaf area dynamics of sugar beet (Beta vulgaris L.", FIELD CROPS RESEARCH, vol. 61, 1999, pages 163 - 177
DOHM JCMINOCHE AEHOLTGRAWE DCAPELLA-GUTIERREZ SZAKRZEWSKI FTAFER HRUPP OSORENSEN TRSTRACKE RREINHARDT R: "The genome of the recently domesticated crop plant sugar beet (Beta vulgaris)", NATURE, vol. 505, no. 7484, 18 December 2013 (2013-12-18), pages 546 - 9, XP055767881, DOI: 10.1038/nature12817
DOHM: "The genome of the recently domesticated crop plant sugar beet (Beta vulgaris", NATURE, vol. 505, no. 7484, 23 January 2014 (2014-01-23), pages 546 - 9, XP055767881, DOI: 10.1038/nature12817
DOLJA, V.V.: "Beet yellows virus: the importance of being different", MOLECULAR PLANT PATHOLOGY, vol. 4, 2003, pages 91 - 98
DOLJA, V.V.KOONIN, E.V.: "The closterovirus-derived gene expression and RNA interference vectors as tools for research and plant biotechnology", FRONTIERS IN MICROBIOLOGY, vol. 4, 2013, pages 83
DUNNINGBYFORD: "Pests, Diseases and Disorders of the Sugar Beet", DELEPLANQUE: BROOMS BARN EXPERIMENTAL STATION, 1982
DUSIPETERS: "Beet mosaic virus: its vector and host relationships", JOURNAL OF PHYTOPATHOLOGY, vol. 147, 1999, pages 293 - 298
FLORES OLVERA H. ET AL.: "Floral and Inflorescence Morphology and Ontogeny in Beta vulgaris, with Special Emphasis on the Ovary Position", ANNALS OF BOTANY, vol. 102, no. 4, 2008, pages 643 - 651
FRANCA ET AL., QUARTERLY REVIEWS OF BIOPHYSICS, vol. 35, 2002, pages 169 - 200
FREE, J. B. ET AL.: "Insect pollination of sugar-beet (Beta vulgaris) seed crops", ANNALS OF APPLIED BIOLOGY, vol. 81, no. 2, 1975, pages 127 - 134
GALLET ET AL.: "Vector-transmission of plant viruses and constraints imposed by virus-vector interactions", CURRENT OPINION IN VIROLOGY, vol. 33, 2018, pages 144 - 150
GARVIN ET AL.: "DEco-TILLING: An inexpensive method for SNP discovery that reduces ascertainment bias", MOLECULAR ECOLOGY NOTES, vol. 7, 2007, pages 735 - 746
GILCHRIST ET AL., MOL. ECOL., vol. 15, 2006, pages 1367 - 1378
GRAYGILDOW: "Luteovirus-aphid interactions", ANNUAL REVIEW OF PHYTOPATHOLOGY, vol. 41, 2003, pages 539 - 566
HALLAUERMIRANDA: "Quantitative Genetics in Maize Breeding", 1981, IOWA STATE UNIVERSITY PRESS
JENSEN: "Plant Breeding Methodology", 1988, JOHN WILEY & SONS, INC.
KOZLOWSKA-MAKULSKA, A. ET AL.: "Aphid transmissibility of different European beet polerovirus isolates", EUROPEAN JOURNAL OF PLANT PATHOLOGY, vol. 125, 2009, pages 337 - 341, XP019728990, DOI: 10.1007/s10658-009-9474-7
LARKIN ET AL., BIOINFORMATICS, vol. 23, 2007, pages 2947 - 294
LEWELLEN, R.T.: "Reaction of sugar beet breeding lines and hybrids to beet chlorosis luteovirus", JOURNAL OF SUGAR BEET RESEARCH, vol. 36, 1999, pages 76
LIMBURG, D. ET AL.: "Characteristics of beet yellows closterovirus transmission to sugar beets by Aphis fabae", PHYTOPATHOLOGY, vol. 87, 1997, pages 766 - 771
LIU ET AL., JOURNAL OF BIOMEDICINE AND BIOTECHNOLOGY, vol. 2012, no. 251364, 2012
LUTERBACHER ET AL.: "Sources of resistance to diseases of sugar beet in related Beta germplasm: I. Foliar diseases", EUPHYTICA, vol. 139, 2004, pages 105 - 121, XP019240772, DOI: 10.1007/s10681-004-2488-5
LUTERBACHER M C ET AL: "Sources of resistance to diseases of sugar beet in related Beta germplasm: II. Soil-borne diseases", EUPHYTICA, KLUWER ACADEMIC PUBLISHERS, DO, vol. 141, no. 1-2, 1 January 2005 (2005-01-01), pages 49 - 63, XP019240828, ISSN: 1573-5060, DOI: 10.1007/S10681-005-5231-Y *
M. K. GRIMMER: "Beet mild yellowing virus resistance derived from wild and cultivated Beta germplasm", PLANT BREEDING, vol. 127, no. 3, 1 June 2008 (2008-06-01), DE, pages 315 - 318, XP093155434, ISSN: 0179-9541, DOI: 10.1111/j.1439-0523.2007.01457.x *
MAKAROVA, K.S., NAT REV MICROBIOL., vol. 9, no. 6, 9 May 2011 (2011-05-09), pages 467 - 477
MARDIS, GENOMICS AND HUMAN GENETICS, vol. 9, 2008, pages 387 - 402
MCGRATH JM ET AL.: "Sugar Beet Breeding", vol. 42, WILEY, article "Plant Breeding Reviews"
MEJLHEDE ET AL., PLANT BREEDING, vol. 125, 2006, pages 461 - 467
MEYERSMILLER, COMPUTER APPLIC. BIOL. SCI., vol. 4, 1988, pages 11 - 17
MINOCHE AEDOHM JCSCHNEIDER JHOLTGRAWE DVIEHOVER PMONTFORT MSORENSEN TRWEISSHAAR BHIMMELBAUER H: "Exploiting single-molecule transcript sequencing for eukaryotic gene prediction", GENOME BIOL., vol. 16, no. 1, 2 September 2015 (2015-09-02), pages 184
MORITANI M ET AL.: "Identification of the predominant nonrestoring allele for Owen-type cytoplasmic male sterility in sugar beet (Beta vulgaris L.): development of molecular markers for the maintainer genotype", MOL BREED., vol. 32, no. 1, June 2013 (2013-06-01), pages 91 - 100
NIETO ET AL., BMC PLANT BIOLOGY, vol. 7, 2007, pages 34 - 42
NOROUZI, P.: "Identification and Confirmation of New Molecular Markers Linked to The Fertility Restorer Genes in Sugar Beet", J. CROP SCI. BIOTECHNOL., vol. 21, 2018, pages 335 - 341, XP036672187, DOI: 10.1007/s12892-018-0078-0
P. HOLLIDAY: "A Dictionary of Plant Pathology", 1992, CAMBRIDGE UNIVERSITY PRESS, pages: 369
PAGAN IGARCIA-ARENAL F: "Tolerance to Plant Pathogens: Theory and Experimental Evidence", INT J MOL SCI., vol. 19, no. 3, 11 March 2018 (2018-03-11), pages 810
PIERIK: "In vitro culture of higher plants", 1999, SPRINGER
R. HOSSAIN ET AL.: "New insights into virus yellows distribution in Europe and effects of beet yellows virus, beet mild yellowing virus, and beet chlorosis virus on sugar beet yield following field inoculation", PLANT PATHOLOGY, vol. 70, no. 3, 29 October 2020 (2020-10-29), pages 584 - 593
RICHARDS: "Plant breeding systems", 1997, TAYLOR & FRANCIS
SALLY A FRANCIS ET AL: "Identification and exploitation of novel disease resistance genes in sugar beet", PEST MANAGEMENT SCIENCE, vol. 59, no. 2, 1 January 2003 (2003-01-01), Hoboken, USA, pages 225 - 230, XP055492871, ISSN: 1526-498X, DOI: 10.1002/ps.569 *
SCHLIEPHAKE, E. ET AL.: "Investigations on the vector transmission of the Beet mild yellowing virus (BMYV) and the Turnip yellows virus (TuYV", JOURNAL OF PLANT DISEASES AND PROTECTION, 2000, pages 81 - 87
SEMAL, J.: "Transmission of Beet mosaic virus from Stellaria media and Capsella bursa-pastoris by Myzus ascalonicus Doncaster", NATURE, vol. 178, 1956, pages 501 - 502
SEYMOUR ET AL., PNAS, vol. 109, 2012, pages 4227 - 4232
SHI ET AL., DIFFERENT MOLECULAR MARKERS TO IDENTIFY THE FERTILITY OF SUGAR BEET MONOGERM GERMPLASM RESOURCES, 2021, Retrieved from the Internet <URL:assets.researchsquare.com/files/rs-230220/v1/327c81e7-6b72-4f2d-9fdc-cb57eae50bOO.pdf?c=1631883847>
SILVA ET AL., CURRENT GENE THERAPY, vol. 11, 2011, pages 11 - 27
SIMMONDS: "Principles of Crop Improvement", LONGMAN GROUP LIMITED, 1979
SRIVASTAVA ET AL.: "Molecular markers, natural history, and evolution", 2004, SINAUER ASSOCIATES
STODDARD ET AL., MOBILE DNA, vol. 5, 2014, pages 7
TAJI ET AL.: "In vitro plant breeding", ROUTLEDGE, 2002, ISBN: 156022908X
ZHANG ET AL., PLANT CELL REP., vol. 12, 2008, pages 1851 - 60
ZHU G ET AL.: "Beta vulgaris", vol. 5, 2003, SCIENCE PRESS/MISSOURI BOTANICAL GARDEN PRESS, article "Magnoliophyta: Caryophyllidae", pages: 354 - 267

Cited By (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
JP7724484B1 (ja) * 2024-03-11 2025-08-18 国立研究開発法人農業・食品産業技術総合研究機構 テンサイ植物の萎黄性ウイルス病抵抗性マーカー及びその利用

Also Published As

Publication number Publication date
EP4658061A1 (fr) 2025-12-10

Similar Documents

Publication Publication Date Title
Veronesi et al. Alfalfa
US11324185B2 (en) Garden bean named HMX0164423
US8609928B2 (en) Squash leaf curl virus (SLCV) resistance in cucurbits
WO2008152134A1 (fr) Plant de poivron amélioré
US20220087187A1 (en) Methods for Producing Corn Plants with Northern Leaf Blight Resistance and Compositions Thereof
US10301641B2 (en) Genetic markers associated with drought tolerance in maize
WO2024161358A1 (fr) Résistance au virus de la jaunisse des betteraves
WO2022208489A1 (fr) Trait d&#39;habitude de croissance semi-déterminé ou déterminé chez cucurbita
US9161501B2 (en) Genetic markers for Orobanche resistance in sunflower
US10041089B1 (en) Resistance alleles in soybean
US12376554B2 (en) Hybrid tomato plant named red coral
US12439884B2 (en) Hybrid tomato plant named HM 8507
US12274233B2 (en) Hybrid tomato plant named HMC44440
US12302854B2 (en) Hybrid tomato plant named HMC44441
US12171185B2 (en) Hybrid tomato plant named HM 7103
US12075747B2 (en) Hybrid tomato plant named HM 8268
US20250113796A1 (en) Cucurbita with determinate growth habit
US12127525B2 (en) Hybrid tomato plant named HM 8237
US11266087B1 (en) Beet cultivar named Amarena
US20240423150A1 (en) Brassica rapa var nipposinica named mazunte
Verma et al. Tree Cotton (Gossypium arboreum L.) Breeding: Current Status and Future Prospects
US20230329182A1 (en) Hybrid tomato rootstock variety setpro
US20250134057A1 (en) Hybrid tomato plant named el rosillo
US20250098626A1 (en) Hybrid tomato plant named curioso
US20250134037A1 (en) Tetraploid watermelon line named w087yr

Legal Events

Date Code Title Description
121 Ep: the epo has been informed by wipo that ep was designated in this application

Ref document number: 24703866

Country of ref document: EP

Kind code of ref document: A1

WWE Wipo information: entry into national phase

Ref document number: 823635

Country of ref document: NZ

WWP Wipo information: published in national office

Ref document number: 823635

Country of ref document: NZ

NENP Non-entry into the national phase

Ref country code: DE

WWP Wipo information: published in national office

Ref document number: 2024703866

Country of ref document: EP