Phloretin glycosyltransferases, polynucleotides encoding these and methods of use
View Patent ↗The invention provides method for producing a plant cell or plant with increased phlorizin or phloretin glycosyltransferase activity, the method comprising transformation of a plant cell or plant with a polynucleotide encoding a polypeptide with phloretin glycosyltransferase activity. The invention also provides host cells, plant cells and plants, genetically modified to contain and or express the polynucleotides.
1. A method for producing a plant cell or plant with at least one of
a) increased phlorizin content, and
b) increased phloretin glycosyltransferase activity,
the method comprising transformation of a plant cell or plant with a polynucleotide encoding a polypeptide with at least 95% sequence identity to the amino acid sequence of any one of SEQ ID NOs: 1 to 5, wherein in order to increase phlorizin content as in a) the plant cell or plant contains phloretin.
2. The method of claim 1 , wherein the polynucleotide encodes a polypeptide with phloretin glycosyltransferase activity.
3. The method of claim 1 , wherein the polynucleotide encodes a polypeptide with the amino acid sequence of SEQ ID NO: 1.
4. The method of claim 1 , wherein the polynucleotide comprises a nucleotide sequence selected from the group consisting of the sequences SEQ ID NOs: 6, 7, 8, 9, 10, 11, 12, 13, 14 and 15, and sequences with at least 95% identity to the nucleotide sequences selected from the group consisting of SEQ ID NOs: 6, 7, 8, 9, 10, 11, 12, 13, 14 and 15.
5. The method of claim 1 , wherein the polynucleotide comprises a nucleotide sequence with at least 95% identity to the sequence of SEQ ID NO: 6.
6. The method of claim 1 , wherein the polynucleotide comprises the nucleotide sequence of SEQ ID NO: 6.
7. The method of claim 1 , wherein the polynucleotide comprises a nucleotide sequence with at least 95% identity to the sequence of SEQ ID NO: 7.
8. The method of claim 1 , wherein the polynucleotide comprises the nucleotide sequence of SEQ ID NO: 7.
9. The method of claim 1 wherein the polynucleotide encodes a polypeptide with the amino acid sequence of any one of SEQ ID NOs: 1 to 5.
10. The method of claim 1 , wherein the polynucleotide comprises a nucleotide sequence selected from the group consisting of the sequences of SEQ ID NOs: 6, 7, 8, 9, 10, 11, 12, 13, 14 and 15.
11. A plant cell or plant produced by the method of claim 1 .
12. A method of producing phlorizin, the method comprising extracting phlorizin from the plant cell or plant of claim 11 .
13. A host cell, plant cell or plant wherein the host cell, plant cell or plant comprises a transgene including a polynucleotide encoding polypeptide comprising the sequence of any one of SEQ ID NOs: 1-5, or a polypeptide with at least 95% identity to the sequence of any one of SEQ ID NOs: 1-5, wherein percent identity is calculated over the entire length of SEQ ID NOs: 1-5.
14. The host cell, plant cell or plant of claim 13 , wherein the polynucleotide encodes a polypeptide with phloretin glycosyltransferase activity.
15. The host cell, plant cell or plant of claim 13 , wherein the polynucleotide encodes a polypeptide comprising a sequence with at least 95% identity to SEQ ID NO: 5.
16. The host cell, plant cell or plant of claim 13 , wherein the polynucleotide encodes a polypeptide comprising the sequence of SEQ ID NO: 5.
17. The hose cell, plant cell or plant of claim 13 comprising the sequence of any one of SEQ ID NOs: 6 to 15, or a sequence with at least 95% identity to the sequence of any one of SEQ ID NOs: 6 to 15, wherein percent identity is calculated over the entire length of SEQ ID NOs: 6 to 15.
18. The host cell, plant cell or plant of claim 17 comprising the sequence of any one of SEQ ID NOs: 6 to 15.
19. The host cell, plant cell or plant of claim 17 comprising the sequence of any one of SEQ ID NOs: 8 to 15, or a sequence with at least 95% identity to the sequence of any one of SEQ ID NOs: 8 to 15, wherein percent identity is calculated over the entire length of SEQ ID NOs: 8 to 15.
20. The host cell, plant cell or plant of claim 19 comprising the sequence of any one of SEQ ID NOs: 8 to 15.
21. The host cell, plant cell or plant of claim 13 , wherein the polynucleotide comprises a sequence with at least 95% sequence identity to the sequence of SEQ ID NO: 14.
22. The host cell, plant cell or plant of claim 13 , wherein the polynucleotide comprises the sequence of SEQ ID NO: 14.
23. The host cell, plant cell or plant of claim 13 , wherein the polynucleotide comprises a sequence with at least 95% sequence identity to the sequence of SEQ ID NO: 15, wherein percent identity is calculated over the entire length of SEQ ID NO: 15.
24. The host cell, plant cell or plant of claim 13 , wherein the polynucleotide comprises the sequence of SEQ ID NO: 15.
25. The host cell, plant cell or plant of claim 13 which comprises a transgene including a polynucleotide encoding a polypeptide comprising the sequence of any one of SEQ ID NOs: 1-5.
26. The host cell, plant cell or plant of claim 13 which comprises a transgene including a polynucleotide encoding a polypeptide comprising the sequence of any one of SEQ ID NOs: 2 to 5, or a polypeptide with at least 95% identity to the sequence of any one of SEQ ID NOs: 2 to 5, wherein percent identity is calculated over the entire length of SEQ ID NOs: 2 to 5.
27. The host cell, plant cell or plant of claim 13 which comprises a transgene including a polynucleotide encoding a polypeptide comprising the sequence of any one of SEQ ID NOs: 2-5.
28. A genetic construct that comprises a polynucleotide encoding a polypeptide comprising the sequence of any one of SEQ ID NOs: 1 to 5, or a polypeptide with at least 95% identity to the sequence of any one of SEQ ID NOs: 1 to 5, wherein percent identity is calculated over the entire length of SEQ ID NOs: 1 to 5; and wherein the polynucleotide is operably linked to a promoter that is at least one of:
a) not normally associated with the polynucleotide in nature,
b) derived from the gene of a plant other than that from which the polynucleotide is derived,
c) derived from a virus,
d) derived from a plant pathogenic bacterium, or
e) derived from a fungus.
29. A host cell comprising the genetic construct of claim 28 .
30. A method for the biosynthesis of phlorizin comprising the steps of culturing a plant host-cell comprising the genetic construct of claim 28 , capable of expressing a phloretin glycosyltransferase, in the presence of a phloretin which may be supplied to, or may be naturally present within the plant host cell.
31. A plant cell which comprises the genetic construct of claim 28 .
32. A plant which comprises the plant cell of claim 31 .