Single nucleotide polymorphism (SNP) markers for stevia

• US 10,370,673 B2
• Filed: 09/25/2015
• Issued: 08/06/2019
• Est. Priority Date: 09/26/2014
• Status: Active Grant

• Current Assignee: PureCircle Sdn Bhd (Ingredion Incorporated)
• Original Assignee: PureCircle Sdn Bhd (Ingredion Incorporated)
• Inventors: Markosyan, Avetik, Ong, Seong Siang, Wong, Yeen Yee, Bu, Yu Cheng, Chen, Jian Ning
• Primary Examiner(s): Stankovic, Bratislav
• Application Number:
  US15/512,271
• Publication Number:
  US 20170283819A1
• Time in Patent Office: 1,411 Days
• Field of Search: None
• CPC Class Codes: A01H 5/12 Leaves
• CPC Class Codes: A23L 27/36 Terpene glycosides
• CPC Class Codes: C07H 21/04 with deoxyribosyl as saccharide radical
• CPC Class Codes: C12N 15/00 Mutation or genetic engineering; DNA or RNA concerning genetic engineering, vectors, e.g. plasmids, or their isolation, preparation or purification; Use of hosts therefor mutants or genetically engineered microorganisms, per se C12N1/00, C12N5/00, C12N7/00; new plants per se A01H; plant reproduction by tissue culture techniques A01H4/00; new animals per se A01K67/00; use of medicinal preparations containing genetic material which is inserted into cells of the living body to treat genetic diseases, gene therapy A61K48/00
• CPC Class Codes: C12N 15/52 Genes encoding for enzymes or proenzymes
• CPC Class Codes: C12N 15/8245 involving modified carbohydrate or sugar alcohol metabolism, e.g. starch biosynthesis
• CPC Class Codes: C12N 5/00 Undifferentiated human, animal or plant cells, e.g. cell lines; Tissues; Cultivation or maintenance thereof; Culture media therefor; plant reproduction by tissue culture techniques A01H4/00
• CPC Class Codes: C12N 5/04 Plant cells or tissues culture media C12N5/0025
• CPC Class Codes: C12Q 1/68 involving nucleic acids
• CPC Class Codes: C12Q 1/6895 for plants, fungi or algae

Abstract

Stevia varieties with high a content of RebD, a high content of RebM, and a high content of RebD and RebM containing various SNP markers and UGT isoforms, are disclosed. Methods of screening for the SNPs are also disclosed as well as for using the SNPs in marker assisted breeding. Further provided are methods for introgressing the disclosed SNPs associated with high RebD and high RebM into stevia plants by selecting plants comprising for one or more SNPs and breeding with such plants to confer such desirable agronomic phenotypes to plant progeny.

Claims

1. A stevia plant comprising eight single nucleotide polymorphisms (SNPs), wherein said SNPs are found in homozygous form, wherein said eight SNPs comprise:

SNP2, which comprises a G to C nucleotide substitution at position number 225 in SEQ ID NO;

1, SNP10, which comprises a G to A nucleotide substitution at position number 187 in SEQ ID NO;

2, SNP12, which comprises a C to T nucleotide exchange at position number 345 in SEQ ID NO;

3, SNP17, which comprises an A to C nucleotide substitution at position number 129 in SEQ ID NO;

4, SNP19, which comprises an A to T nucleotide substitution at position number 173 in SEQ ID NO;

5, SNP20, which comprises a G to A nucleotide substitution at position number 221 in SEQ ID NO;

6, SNP22, which comprises a T to G nucleotide substitution at position number 160 in SEQ ID NO;

7, and SNP24, which comprises a G to A nucleotide substitution at position number 325 in SEQ ID NO;

8.

2. The stevia plant of claim 1, wherein the leaves of said stevia plant have a rebaudioside D content of between 0.6% and 3.3% of dry weight.

3. The stevia plant of claim 1, wherein the leaves of said stevia plant have a rebaudioside M content of between 0.5% and 1.15% of dry weight.

4. The stevia plant of claim 1, wherein the leaves of said stevia plant have a rebaudioside D content of between 0.6% and 3.3% of dry weight, and a rebaudioside M content of between 0.5% and 1.15% of dry weight.

5. The stevia plant of claim 1, further comprising isoforms of UDP-glucosyltransferase 76G1 (UGT76G1), wherein said isoforms correspond to SEQ ID NOs:

19-23, and isoforms of UDP-glucosyltransferase 91D2 (UGT91D2), wherein said isoforms correspond to SEQ ID NOs;

24-54, and wherein said stevia plant has high rebaudioside D content.

6. The stevia plant of claim 1, further comprising isoforms of UDP-glucosyltransferase 76G1 (UGT76G1), wherein said isoforms correspond to SEQ ID NOs:

9-13, and isoforms of UDP-glucosyltransferase 91D2 (UGT91D2), wherein said isoforms correspond to SEQ ID NOs;

86-116, and wherein said stevia plant has high rebaudioside M content.

7. The stevia plant of claim 1, further comprising isoforms of UDP-glucosyltransferase 76G1 (UGT76G1), wherein said isoforms correspond to SEQ ID NOs:

14-18, and isoforms of UDP-glucosyltransferase 91D2 (UGT91D2), wherein said isoforms correspond to SEQ ID NOs;

55-85, and wherein said stevia plant has high rebaudioside M content.

8. The stevia plant of claim 6, wherein said stevia plant further comprises a high rebaudioside D content.

9. The stevia plant of claim 7, wherein said stevia plant further comprises a high rebaudioside D content.

10. A method of producing a commodity plant product, comprising obtaining the plant of claim 1, or a part thereof, and producing the commodity plant product from the plant or part thereof, wherein the commodity plant product is a composition of glycosides.