Production of hydroxylated fatty acids in genetically modified plants

Abstract

This invention relates to plant fatty acid hydroxylases. Methods to use conserved amino acid or nucleotide sequences to obtain plant fatty acid hydroxylases are described. As described is the use of cDNA clones encoding a plant hydroxylase to produce a family of hydroxylated fatty acids in transgenic plants. In addition, the use of genes encoding fatty acid hydroxylases or desaturases to alter the level of lipid fatty acid unsaturation in transgenic plants is described.

What is claimed is:  
     
         1 . A method of altering an amount of an unsaturated fatty acid in a seed of a plant comprising: decreasing a fatty acid desaturase activity in the seed by genetic manipulation of at least one of fatty acid desaturase or fatty acid hydroxylase.  
     
     
         2 . The method of  claim 1 , wherein an endogenous gene for said fatty acid hydroxylase is mutated and thereby decreases fatty acid hydroxylase activity in the seed.  
     
     
         3 . The method of  claim 1 , wherein said plant is transformed with a nucleic acid containing a sequence which encodes a fatty acid hydroxylase or derivative thereof.  
     
     
         4 . The method of  claim 3 , wherein said derivative is a dominant negative mutant which thereby alters the amount of the unsaturated fatty acid in the seed.  
     
     
         5 . The method of  claim 3 , wherein said derivative is a mutant fatty acid hydroxylase in which one or more essential histidine residues have been mutated which thereby alters the amount of the unsaturated fatty acid in the seed.  
     
     
         6 . The method of  claim 1 , wherein an endogenous gene for said fatty acid desaturase is mutated and thereby decreases fatty acid desaturase activity in the seed.  
     
     
         7 . The method of  claim 1 , wherein said plant is transformed with a nucleic acid containing a sequence which encodes a fatty acid desaturase or derivative thereof.  
     
     
         8 . The method of  claim 7 , wherein said derivative is a dominant negative mutant which thereby alters the amount of the unsaturated fatty acid in the seed.  
     
     
         9 . The method of  claim 7 , wherein said derivative is a mutant fatty acid desaturase in which one or more essential histidine residues have been mutated which thereby alters the amount of the unsaturated fatty acid in the seed.  
     
     
         10 . The method of  claim 1 , wherein said plant is selected from the group consisting of rapeseed,  Crambe, Brassica juncea , canola, flax, sunflower, safflower, cotton, cuphea, soybean, peanut, coconut, oil palm and corn.  
     
     
         11 . A method of altering an amount of a unsaturated fatty acid comprising: 
 (a) transforming a plant cell with a nucleic acid containing a sequence which encodes a fatty acid hydroxylase or a dominant negative mutant of fatty acid hydroxylase or a dominant negative mutant of fatty acid desaturase,    (b) growing a seed-bearing plant from the transformed plant cell of step (a), and    (c) identifying a seed from the plant of step (b) with the altered amount of the unsaturated fatty acid in the seed.    
     
     
         12 . The method of  claim 11 , wherein said nucleic acid contains a sequence which encodes the dominant negative mutant of fatty acid hydroxylase in which one or more essential histidine residues have been mutated.  
     
     
         13 . The method of  claim 11 , wherein said nucleic acid contains a sequence which encodes the dominant negative mutant of fatty acid hydroxylase which thereby alters the amount of the unsaturated fatty acid in the seed.  
     
     
         14 . The method of  claim 11 , wherein said nucleic acid contains a sequence which encodes the dominant negative mutant of fatty acid desaturase in which one or more essential histidine residues have been mutated.  
     
     
         15 . The method of  claim 11 , wherein said nucleic acid contains a sequence which encodes the dominant negative mutant of fatty acid desaturase which thereby alters the amount of the unsaturated fatty acid in the seed.  
     
     
         16 . The method of  claim 11 , wherein said plant is selected from the group consisting of rapeseed,  Crambe, Brassica juncea , canola, flax, sunflower, safflower, cotton, cuphea, soybean, peanut, coconut, oil palm and corn.  
     
     
         17 . A recombinant nucleic acid suitable for use in  claim 1 , wherein said nucleic acid contains a sequence encoding a fatty acid hydroxylase with an amino acid identity of 60% or greater to SEQ ID NO:4.  
     
     
         18 . The recombinant nucleic acid of  claim 17 , wherein the amino acid identity is 90% or greater to SEQ ID NO:4.  
     
     
         19 . The recombinant nucleic acid of  claim 17 , wherein the amino acid identity is 100% of SEQ ID NO:4.  
     
     
         20 . The recombinant nucleic acid of  claim 17 , wherein said nucleic acid contains a sequence having a nucleotide identity of 90% or greater to SEQ ID NO:1, 2 or 3.  
     
     
         21 . The recombinant nucleic acid of  claim 17 , wherein said nucleic acid contains SEQ ID NO:1, 2 or 3.  
     
     
         22 . The recombinant nucleic acid of  claim 17 , wherein said sequence is obtainable from a plant species producing a hydroxylated fatty acid.  
     
     
         23 . A recombinant nucleic acid suitable for use in  claim 1 , wherein said nucleic acid contains a sequence encoding at least one of fatty acid desaturase or fatty acid hydroxylase.  
     
     
         24 . The recombinant nucleic acid of  claim 23 , wherein said sequence is obtainable from  Ricinus communis  (L.) (castor).  
     
     
         25 . The recombinant nucleic acid of  claim 23 , wherein said sequence is obtainable from  Lesquerella fendleri.    
     
     
         26 . The recombinant nucleic acid of  claim 23 , wherein said nucleic acid contains a sequence encoding at least one of fatty acid desaturase or fatty acid hydroxylase in which one or more essential histidine residues have been mutated.  
     
     
         27 . The method of  claim 1  further comprising: processing the seed containing the altered amount of the unsaturated fatty acid to obtain oil and/or seed meal.  
     
     
         28 . Oil obtained by the method of  claim 27 .  
     
     
         29 . Seed meal obtained by the method of  claim 27 .  
     
     
         30 . Plant obtained by the method of  claim 1 .  
     
     
         31 . The method of  claim 11  further comprising: processing the seed containing the altered amount of the unsaturated fatty acid to obtain oil and/or seed meal.  
     
     
         32 . Oil obtained by the method of  claim 31 .  
     
     
         33 . Seed meal obtained by the method of  claim 31 .  
     
     
         34 . Plant obtained by the method of  claim 11.