Fatty acid desaturases and elongases and uses thereof

Patent metadata

Field	Value
Publication number	US-9493520-B2
Application number	US-201013384277-A
Country	US
Kind code	B2
Filing date	Jul 15, 2010
Priority date	Jul 17, 2009
Publication date	Nov 15, 2016
Grant date	Nov 15, 2016

Abstract

The invention provides isolated nucleic acid molecules which encode novel fatty acid desaturases and elongases from the organism Emiliana huxleyi . The invention also provides recombinant expression vectors containing desaturase or elongase nucleic acid molecules, host cells into which the expression vectors have been introduced, and methods for large-scale production of long chain polyunsaturated fatty acids (LCPUFAs), e.g. arachidonic acid (ARA), eicosapentaenoic acid (EPA) or docosahexaenoic acid (DHA).

First claim

The invention claimed is:

1. A polynucleotide comprising an expression control sequence operatively linked to a first nucleic acid sequence selected from the group consisting of:
a) the nucleic acid sequence of SEQ ID NO: 3;
b) a nucleic acid sequence encoding a polypeptide comprising the amino acid sequence of SEQ ID NO: 4;
c) a nucleic acid sequence having at least 70% sequence identity to the nucleic acid sequence of a), wherein said nucleic acid sequence encodes a polypeptide having elongase activity;
d) a nucleic acid sequence encoding a polypeptide having elongase activity and comprising an amino acid sequence which has at least 82% sequence identity to the amino acid sequence of SEQ ID NO: 4; and
e) a nucleic acid sequence which is capable of hybridizing under stringent conditions comprising hybridization in 6× sodium chloride/sodium citrate (SSC) at approximately 45° C. followed by one or more wash steps in 0.2× SSC, 0.1% SDS at 50 to 65° C. to the nucleic acid sequence of a), wherein said nucleic acid sequence encodes a polypeptide having elongase activity,
wherein said expression control sequence is heterologous to said first nucleic acid sequence,
and wherein said polynucleotide further comprises a second nucleic acid sequence selected from the group consisting of:
i) the nucleic acid sequence of SEQ ID NO: 9;
ii) a nucleic acid sequence encoding a polypeptide comprising the amino acid sequence of SEQ ID NO: 10;
iii) a nucleic acid sequence having at least 70% sequence identity to the nucleic acid sequence of i), wherein said nucleic acid sequence encodes a polypeptide having desaturase activity;
iv) a nucleic acid sequence encoding a polypeptide having desaturase activity and comprising an amino acid sequence which has at least 82% sequence identity to the amino acid sequence of SEQ ID NO: 10; and
v) a nucleic acid sequence which is capable of hybridizing under stringent conditions comprising hybridization in 6× sodium chloride/sodium citrate (SSC) at approximately 45° C. followed by one or more wash steps in 0.2× SSC, 0.1% SDS at 50 to 65° C. to the nucleic acid sequence of i), wherein said nucleic acid sequence encodes a polypeptide having desaturase activity.

2. The polynucleotide of claim 1 , wherein the polynucleotide further comprises a terminator sequence operatively linked to the first nucleic acid sequence.

3. A vector comprising the polynucleotide of claim 1 .

4. A host cell comprising:
a) the polynucleotide of claim 1 ; or
b) a vector comprising said polynucleotide.

5. A method for the manufacture of a polyunsaturated fatty acid, comprising:
a) cultivating the host cell of claim 4 under conditions which allow for the production of a polyunsaturated fatty acid in said host cell; and
b) obtaining said polyunsaturated fatty acid from said host cell.

6. The method of claim 5 , wherein said polyunsaturated fatty acid is arachidonic acid (ARA), eicosapentaenoic acid (EPA), and/or docosahexaenoic acid (DHA).

7. A method for the manufacture of an oil, lipid, or fatty acid composition, comprising:
a) cultivating the host cell of claim 4 under conditions which allow for the production of a polyunsaturated fatty acid in said host cell;
b) obtaining said polyunsaturated fatty acid from said host cell; and
c) formulating the polyunsaturated fatty acid as an oil, lipid, or fatty acid composition.

8. The method of claim 7 , wherein the oil, lipid, or fatty acid composition is used for feed, foodstuffs, cosmetics, or medicaments.

9. A method for the manufacture of a polypeptide, comprising:
a) cultivating a host cell comprising the polynucleotide of claim 1 or a vector comprising said polynucleotide under conditions which allow for the production of a polypeptide encoded by the first nucleic acid sequence or the second nucleic acid sequence; and
b) obtaining the polypeptide from the host cell of step a).

10. A non-human transgenic organism comprising:
a) the polynucleotide of claim 1 ; or
b) a vector comprising said polynucleotide.

11. The non-human transgenic organism of claim 10 , which is a plant, plant part, or plant seed.

12. A method for the manufacture of a polyunsaturated fatty acid, comprising:
a) cultivating the non-human transgenic organism of claim 10 under conditions which allow for the production of a polyunsaturated fatty acid in said non-human transgenic organism; and
b) obtaining said polyunsaturated fatty acid from said non-human transgenic organism.

13. A method for the manufacture of polyunsaturated fatty acids, comprising:
a) cultivating a plant comprising the polynucleotide of claim 1 or a vector comprising said polynucleotide under conditions which allow for the production of polyunsaturated fatty acids in said plant or seeds thereof; and
b) obtaining said polyunsaturated fatty acids from said plant or seeds thereof.

14. A method for the manufacture of an oil-, lipid- or fatty acid-composition, comprising:
a) cultivating a plant comprising the polynucleotide of claim 1 or a vector comprising said polynucleotide under conditions which allow for the production of polyunsaturated fatty acids in said plant or seeds thereof; and
b) obtaining an oil-, lipid- or fatty acid-composition from said plant or seeds thereof.

15. A method for the manufacture of polyunsaturated fatty acids, comprising:
a) cultivating a plant comprising a first nucleic acid sequence encoding a polypeptide having elongase activity and a second nucleic acid sequence encoding a polypeptide having desaturase activity under conditions which allow for the production of polyunsaturated fatty acids in said plant or seeds thereof; and
b) obtaining said polyunsaturated fatty acids from said plant or seeds thereof,
wherein said first nucleic acid sequence is selected from the group consisting of:
i) the nucleic acid sequence of SEQ ID NO: 3;
ii) a nucleic acid sequence encoding a polypeptide comprising the amino acid sequence of SEQ ID NO: 4;
iii) a nucleic acid sequence having at least 70% sequence identity to the nucleic acid sequence of i);
iv) a nucleic acid sequence encoding a polypeptide having at least 82% sequence identity to the amino acid sequence of SEQ ID NO: 4; and
v) a nucleic acid sequence which is capable of hybridizing under stringent conditions comprising hybridization in 6× sodium chloride/sodium citrate (SSC) at approximately 45° C. followed by one or more wash steps in 0.2× SSC, 0.1% SDS at 50 to 65° C. to the nucleic acid sequence of i),
and wherein said second nucleic acid sequence is selected from the group consisting of:
i) the nucleic acid sequence of SEQ ID NO: 9;
ii) a nucleic acid sequence encoding a polypeptide comprising the amino acid sequence of SEQ ID NO: 10;
iii) a nucleic acid sequence having at least 70% sequence identity to the nucleic acid sequence of i);
iv) a nucleic acid sequence encoding a polypeptide having at least 82% sequence identity to the amino acid sequence of SEQ ID NO: 10; and
v) a nucleic acid sequence which is capable of hybridizing under stringent conditions comprising hybridization in 6× sodium chloride/sodium citrate (SSC) at approximately 45° C. followed by one or more wash steps in 0.2× SSC, 0.1% SDS at 50 to 65° C. to the nucleic acid sequence of i).

16. The method of claim 15 , wherein the polyunsaturated fatty acids are obtained from the seeds of said plant.

17. The method of claim 15 , comprising obtaining an oil-, lipid- or fatty acid-composition from said plant or seeds thereof, and obtaining the polyunsaturated fatty acids from said oil-, lipid- or fatty

Assignees

• Bauer Jörg
• Napier Johnathan A
• Sayanova Olga
• Basf Plant Science Co Gmbh

Inventors

• Bauer Jörg
• Napier Johnathan A
• Sayanova Olga

Classifications

• C12N9/1029

  transferring groups other than amino-acyl groups (2.3.1) · CPC title

• C12N9/0083

  Miscellaneous (1.14.99) · CPC title

• C07K14/435 Primary

  from animals; from humans · CPC title

• C12Y602/01003

  Long-chain-fatty-acid-CoA ligase (6.2.1.3) · CPC title

• C12Y114/19

  with oxidation of a pair of donors resulting in the reduction of molecular oxygen to two molecules of water (1.14.19) · CPC title