Sucrose transporter genes for increasing plant seed lipids
This invention relates to polynucleotide sequences encoding SUT2 or SUT4 sucrose transporter genes. Methods for increasing seed oil content and evaluating increased oil content in a plant seed are described. The compositions and methods disclosed herein employ a variety of sequences that encode sucrose transporters and a variety of sequences that influence fatty acid accumulation, including for example, DGAT, Lec1 and ODP1 transcription factor. In specific embodiments, overexpression of SUT2 and/or SUT4 sucrose transporters in combination with DGAT genes further increase plant seed oil production compared to a high oil plant comprising recombinant DNA constructs that do not overexpress SUT2 or SUT4 transporters.
1. A soybean plant seed with increased oil content comprising in its genome:
(a) a first recombinant DNA construct comprising a first polynucleotide operably linked to at least one first regulatory element, wherein said first polynucleotide comprises a diacylglycerol acyltransferase (DGAT) coding region encoding a polypeptide having at least 95% sequence identity to SEQ ID NO:180, based on the Clustal W method of alignment with pairwise alignment default parameters of KTUPLE=1, GAP PENALTY=3, WINDOW=5 and DIAGONALS SAVED=5, wherein said DGAT polypeptide has DGAT activity;
(b) a second recombinant DNA construct comprising a second polynucleotide operably linked to at least one second regulatory element, wherein said second polynucleotide encodes a SUT4 sucrose transporter polypeptide, wherein said SUT4 sucrose transporter polypeptide has at least 95% sequence identity to SEQ ID NO:38, based on the Clustal W method of alignment with pairwise alignment default parameters of KTUPLE=1, GAP PENALTY=3, WINDOW=5 and DIAGONALS SAVED=5, and wherein said SUT4 sucrose transporter polypeptide has sucrose transport activity; and
(c) wherein said soybean plant seed exhibits increased oil content as a percentage of seed weight when compared to a control soybean plant seed comprising said first recombinant DNA construct but not comprising said second recombinant DNA construct.
2. The plant seed of claim 1 , wherein the first polynucleotide encodes a polypeptide having the amino acid sequence of SEQ ID NO:180, 184, 186, 188, or 190.
3. The plant seed of claim 1 , wherein the second polynucleotide encodes a polypeptide having the amino acid sequence of SEQ ID NO:38.
4. A soybean plant grown from the seed of claim 1 .
5. A method of increasing oil content in a soybean plant seed comprising:
(a) introducing into a regenerable soybean plant cell:
(i) a first recombinant DNA construct comprising a first polynucleotide encoding a DGAT polypeptide having at least 95% sequence identity to SEQ ID NO:180, based on the Clustal W method of alignment with pairwise alignment default parameters of KTUPLE=1, GAP PENALTY=3, WINDOW=5 and DIAGONALS SAVED=5, and
(ii) a second recombinant DNA construct comprising a second polynucleotide encoding a SUT4 sucrose transporter polypeptide, wherein said SUT4 sucrose transporter polypeptide has at least 95% sequence identity to SEQ ID NO:38, based on the Clustal W method of alignment with pairwise default parameters of KTUPLE=1, GAP PENALTY=3, WINDOW=5 and DIAGONALS SAVED=5, and wherein said SUT4 sucrose transporter polypeptide has sucrose transport activity;
(b) regenerating a transgenic plant from the regenerable plant cell after step (a), wherein the transgenic plant comprises in its genome said first recombinant DNA construct and said second recombinant DNA construct; and,
(c) obtaining a progeny plant derived from the transgenic plant of step (b), wherein said progeny plant comprises in its genome said first recombinant DNA construct and said second recombinant DNA construct, wherein said progeny plant exhibits increased oil content as a percentage of seed weight when compared to a control soybean plant comprising said first recombinant DNA construct but not comprising said second recombinant DNA construct.
6. The method of claim 5 , wherein the second polynucleotide encodes a polypeptide having the amino acid sequence of SEQ ID NO:38.
7. The method of claim 5 , wherein the first polynucleotide encodes a polypeptide having the amino acid sequence of SEQ ID NO:180, 184, 186, 188, or 190.
8. A method of evaluating increased seed oil content in a soybean plant seed comprising:
evaluating seed from a progeny plant derived from a transgenic plant for increased oil content, the transgenic plant comprising in its genome:
(i) a first recombinant DNA construct comprising a first polynucleotide encoding a DGAT polypeptide having at least 95% sequence identity to SEQ ID NO:180, based on the Clustal W method of alignment with pairwise alignment default parameters of KTUPLE=1, GAP PENALTY=3, WINDOW=5 and DIAGONALS SAVED=5, wherein said DGAT polypeptide has DGAT activity; and
(ii) a second recombinant DNA construct comprising a second polynucleotide encoding a SUT4 sucrose transporter polypeptide, wherein said SUT4 sucrose transporter polypeptide has at least 95% sequence identity to SEQ ID NO:38, based on the Clustal W method of alignment with pairwise alignment default parameters of KTUPLE=1, GAP PENALTY=3, WINDOW=5 and DIAGONALS SAVED=5, and wherein said SUT4 sucrose transporter polypeptide has sucrose transport activity;
wherein the progeny plant comprises in its genome said first recombinant DNA construct and said second recombinant DNA construct;
and wherein the seed from the progeny plant shows-increased oil content compared to a control soybean plant not comprising said first recombinant DNA construct and said second recombinant DNA construct.
9. The method of claim 8 , wherein the second polynucleotide encodes a polypeptide having an amino acid sequence of SEQ ID NO: 38.
10. The method of claim 8 , wherein the first polynucleotide encodes a polypeptide having the amino acid sequence of SEQ ID NO: 180, 184, 186, 188, or 190.
11. The plant seed of claim 1 , wherein said first regulatory element comprises a seed-specific promoter, and wherein said second regulatory element comprises a different seed-specific promoter.
12. The plant seed of claim 11 , wherein the first seed-specific promoter, the second seed-specific promoter or a combination thereof is selected from the group consisting of the alpha prime subunit of beta conglycinin promoter, soybean sucrose synthase promoter, kunitx trypsin inhibitor 3 promoter, annexin promoter, Glyl promoter, beta subunit of beta conglycinin promoter, P34/Gly Bd m 30 K promoter, albumin promoter, Leg A1 promoter and Leg A2 promoter.
13. The plant seed of claim 12 , wherein both the first and the second seed specific promoters are selected from the group.
14. The plant seed of claim 1 , wherein the plant seed exhibits an overall increased oil content of 1% or greater when compared to the control plant.
15. The plant seed of claim 1 , wherein the plant seed exhibits an overall increased oil content of 5% or greater when compared to the control plant.
16. The plant seed of claim 1 , wherein the plant seed exhibits an overall increased oil content of 10% or greater when compared to the control plant.
17. The method of claim 5 , wherein said first regulatory sequence comprises a seed-specific promoter, and wherein said second regulatory sequence comprises a different seed-specific promoter.
18. The method of claim 5 , wherein the plant seed exhibits an overall increased oil content of 1% or greater when compared to control plant.
19. The method of claim 5 , wherein the plant seed exhibits an overall increased oil content of 5% or greater when compared to the control plant.
20. The method of claim 5 , wherein the plant seed exhibits an overall increased oil content of 5% or greater when compared to the control plant.