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US 5,670,366 AGranted
Recombinant DNA sequences encoding phospholipase
AssigneeKansas State University Research Foundation
InventorXuemin Wang
Life of this patent
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Abstract
Substantially isolated and purified eukaryotic nucleotide sequences encoding for plant phospholipases (e.g., phospholipase D) are disclosed which can be inserted into an expression vector and used for the recombinant expression of substantially purified phospholipases. An exemplary phospholipase D cDNA clone contains a 2424-bp open reading frame encompassing 808 amino acids.
Claims24
A recombinant DNA sequence,
- wherein the DNA sequence encodes a phospholipase D, and hybridizes to an oligonucleotide when the DNA sequence and the oligonucleotide are held at 59.degree. C. for 18 hours in a solution including 6X SSC, 10 mM (Na).sub.3 PO.sub.4, 1 mM EDTA, 0.5% SDS, 0.1% nonfat dried milk, and 50 .mu.g/ml denatured salmon sperm DNA,
- wherein the oligonucleotide is selected from the group consisting of GARGARACNGTNGGNTTYGGNAARGGNGT (SEQ ID #3) and YTNTAYGCNACNATNGAKYT (SEQ ID #4), wherein N is inosine, R is A or G, Y is T or C, and K is T or G.
A vector comprising a recombinant DNA sequence,
- wherein the DNA sequence encodes a phospholipase D, and hybridizes to an oligonucleotide when the DNA sequence and the oligonucleotide are held at 59.degree. C. for 18 hours in a solution including 6X SSC, 10 mM (Na).sub.3 PO.sub.4, 1 mM EDTA, 0.5% SDS, 0.1% nonfat dried milk, and 50 .mu.g/ml denatured salmon sperm DNA,
- wherein the oligonucleotide is selected from the group consisting of GARGARACNGTNGGNTTYGGNAARGGNGT (SEQ ID #3) and YTNTAYGCNACNATNGAKYT (SEQ ID #4), wherein N is inosine, R is A or G, Y is T or C, and K is T or G.
A cell comprising a recombinant DNA sequence,
- wherein the DNA sequence encodes a phospholipase D, and hybridizes to an oligonucleotide when the DNA sequence and the oligonucleotide are held at 59.degree. C. for 18 hours in a solution including 6X SSC, 10 mM (Na).sub.3 PO.sub.4, 1 mM EDTA, 0.5% SDS, 0.1% nonfat dried milk, and 50 .mu.g/ml denatured salmon sperm DNA,
- wherein the oligonucleotide is selected from the group consisting of GARGARACNGTNGGNTTYGGNAARGGNGT (SEQ ID #3) and YTNTAYGCNACNATNGAKYT (SEQ ID #4), wherein N is inosine, R is A or G, Y is T or C, and K is T or G.
Dependencies are inferred from the claim text.