Method for increasing the omega-hydroxylase activity in candida tropicals
AssigneeHenkel Corporation
InventorsStephen Picataggio, Tracy Rohrer, L. Dudley Eirich
Life of this patent
Estimating the term…
Abstract
The omega-hydroxylase activity in the yeast Candida tropicalis is increased by amplification of the P450ALK and P450RED genes. The specific productivity (grams of dicarboxylic acid/liter/hr) of substantially pure alpha, omega-dicarboxylic acids is increased by culturing C. tropicalis strains having amplified P450ALK and P450RED genes.
Claims74
A process for increasing the omega-hydroxylase activity in C. tropicalis comprising introducing into the genome of said C. tropicalis one or more copies of a P450ALK or P450RED gene or a combination thereof.
A transformed C. tropicalis host having an increased copy number of the P450ALK gene relative to the wild type strain wherein the chromosomal POX4A gene in said C. tropicalis host is disrupted through homologous recombination of said POX4A gene with a linear DNA fragment comprised of a URA3A selectable marker gene wherein said selectable marker gene is flanked on both ends by DNA sequences having homology to said POX4A gene.
A transformed C. tropicalis host having an increased copy number of the P450RED gene relative to the wild type strain wherein the chromosomal POX4A gene in said C. tropicalis host is disrupted through homologous recombination of said POX4A gene with a linear DNA fragment comprised of a URA3A selectable marker gene wherein said selectable marker gene is flanked on both ends by DNA sequences having homology to said POX4A gene.
A transformed C. tropicalis host having an increased copy number of both the P450ALK and P450RED genes relative to the wild type strain wherein the chromosomal POX4A gene in said C. tropicalis host is disrupted through homologous recombination of said POX4A gene with a linear DNA fragment comprised of a URA3A selectable marker gene wherein said selectable marker gene is flanked on both ends by DNA sequences having homology to said POX4A gene.
A transformed C. tropicalis host having an increased copy number of the P450ALK gene relative to the wild type strain wherein the chromosomal POX4B gene in said C. tropicalis host is disrupted through homologous recombination of said POX4B gene with a linear DNA fragment comprised of a URA3A selectable marker gene wherein said selectable marker gene is flanked on both ends by DNA sequences having homology to said POX4B gene.
A transformed C. tropicalis host having an increased copy number of the P450RED gene relative to the wild type strain wherein the chromosomal POX4B gene in said C. tropicalis host is disrupted through homologous recombination of said POX4B gene with a linear DNA fragment comprised of a URA3A selectable marker gene wherein said selectable marker gene is flanked on both ends by DNA sequences having homology to said POX4B gene.
A transformed C. tropicalis host having an increased copy number of both the P450ALK and P450RED genes relative to the wild type strain wherein the chromosomal POX4B gene in said C. tropicalis host is disrupted through homologous recombination of said POX4B gene with a linear DNA fragment comprised of a URA3A selectable marker gene wherein said selectable marker gene is flanked on both ends by DNA sequences having homology to said POX4B gene.
A transformed C. tropicalis host having an increasd copy number of the P450ALK gene relative to the wild type strain wherein one of the chromosomal POX5 genes in said C. tropicalis host is disrupted through homologous recombination of said POX5 gene with a linear DNA fragment comprised of a URA3A selectable marker gene wherein said selectable marker gene is flanked on both ends by DNA sequences having homology to said POX5 gene.
A transformed C. tropicalis host having an increased copy number of the P450RED gene relative to the wild type strain wherein one of the chromosomal POX5 genes in said C. tropicalis host is disrupted through homologous recombination of said POX5 gene with a linear DNA fragment comprised of a URA3A selectable marker gene wherein said selectable marker gene is flanked on both ends by DNA sequences having homology to said POX5 gene.
A transformed C. tropicalis host having an increased copy number of both the P450ALK and P450RED genes relative to the wild type strain wherein one of the chromosomal POX5 genes in said C. tropicalis host is disrupted through homologous recombination of said POX5 gene with a linear DNA fragment comprised of a URA3A selectable marker gene wherein said selectable marker gene is flanked on both ends by DNA sequences having homology to said POX5 gene.
A transformed C. tropicalis host having an increased copy number of the P450ALK gene relative to the wild type strain wherein the chromosomal POX4A and one of the chromosomal POX5 genes in said C. tropicalis host are disrupted through homologous recombination of each of said POX4A and POX5 genes with a linear DNA fragment comprised of a URA3A selectable marker gene wherein said selectable marker gene is flanked on both ends by DNA sequences having homology to said POX4A or one of said POX5 genes respectively.
A transformed C. tropicalis host having an increased copy number of the P450RED gene relative to the wild type strain wherein the chromosomal POX4A and one of the chromosomal POX5 genes in said C. tropicalis host are disrupted through homologous recombination of each of said POX4A and POX5 genes with a linear DNA fragment comprised of a URA3A selectable marker gene wherein said selectable marker gene is flanked on both ends by DNA sequences having homology to said POX4A or one of said POX5 genes respectively.
A transformed C. tropicalis host having an increased copy number of both the P450ALK and P450RED genes relative to the wild type strain wherein the chromosomal POX4A and one of the chromosomal POX5 genes in said C. tropicalis host are disrupted through homologous recombination of each of said POX4A and POX5 genes with a linear DNA fragment comprised of a URA3A selectable marker gene wherein said selectable marker gene is flanked on both ends by DNA sequences having homology to said POX4A or one of said POX5 genes respectively.
A transformed C. tropicalis host having an increased copy number of the P450ALK gene relative to the wild type strain wherein the chromosomal POX4A and POX4B genes in said C. tropicalis host are disrupted through homologous recombination of each of said POX4A and POX4B genes with a linear DNA fragment comprised of a URA3A selectable marker gene wherein said selectable marker gene is flanked on both ends by DNA sequences having homology to said POX4A or POX4B gene respectively.
A transformed C. tropicalis host having an increased copy number of the P450RED gene relative to the wild type strain wherein the chromosomal POX4A and POX4B genes in said C. tropicalis host are disrupted through homologous recombination of each of said POX4A and POX4B genes with a linear DNA fragment comprised of a URA3A selectable marker gene wherein said selectable marker gene is flanked on both ends by DNA sequences having homology to said POX4A or POX4B gene respectively.
A transformed C. tropicalis host having an increased copy number of the P450ALK and P450RED gene relative to the wild type strain wherein the chromosomal POX4A and POX4B genes in said C. tropicalis host are disrupted through homologous recombination of each of said POX4A and POX4B genes with a linear DNA fragment comprised of a URA3A selectable marker gene wherein said selectable marker gene is flanked on both ends by DNA sequences having homology to said POX4A or POX4B gene respectively.
A transformed C. tropicalis host having an increased copy number of the P450ALK gene relative to the wild type strain wherein both copies of the chromosomal POX5 genes in said C. tropicalis host are disrupted through homologous recombination of each of said POX5 genes with a linear DNA fragment comprised of a URA3A selectable marker gene wherein said selectable marker gene is flanked on both ends by DNA sequences having homology to both of said POX5 genes.
A transformed C. tropicalis host having an increased copy number of the P450RED gene relative to the wild type strain wherein both copies of the chromosomal POX5 genes in said C. tropicalis host are disrupted through homologous recombination of each of said POX5 genes with a linear DNA fragment comprised of a URA3A selectable marker gene wherein said selectable marker gene is flanked on both ends by DNA sequences having homology to both of said POX5 genes.
A transformed C. tropicalis host having an increased copy number of the P450ALK and P450RED genes relative to the wild type strain wherein both copies of the chromosomal POX5 genes in said C. tropicalis host are disrupted through homologous recombination of each of said POX5 genes with a linear DNA fragment comprised of a URA3A selectable marker gene wherein said selectable marker gene is flanked on both ends by DNA sequences having homology to both of said POX5 genes.
A transformed C. tropicalis host having an increased copy number of the P450ALK gene relative to the wild type strain wherein both copies of the chromosomal POX5 gene and the POX4A gene in said C. tropicalis host are disrupted through homologous recombination of each of both of said POX5 genes and said POX4A gene with a linear DNA fragment comprised of a URA3A selectable marker gene wherein said selectable marker gene is flanked on both ends by DNA sequences having homology to both of said POX5 genes or said POX4A gene respectively.
A transformed C. tropicalis host having an increased copy number of the P450RED gene relative to the wild type strain wherein both copies of the chromosomal POX5 genes and the POX4A gene in said C. tropicalis host are disrupted through homologous recombination of each of both of said POX5 genes and said POX4A gene with a linear DNA fragment comprised of a URA3A selectable marker gene wherein said selectable marker gene is flanked on both ends by DNA sequences having homology to both of said POX5 genes or said POX4A gene respectively.
A transformed C. tropicalis host having an increased copy number of both the P450ALK and P450RED genes relative to the wild type strain wherein both copies of the chromosomal POX5 gene and the POX4A gene in said C. tropicalis host are disrupted through homologous recombination of each of both of said POX5 genes and said POX4A gene with a linear DNA fragment comprised of a URA3A selectable marker gene wherein said selectable marker gene is flanked on both ends by DNA sequences having homology to both of said POX5 genes or said POX4A gene respectively.
A transformed C. tropicalis host having an increased copy number of the P450ALK gene relative to the wild type strain wherein both copies of the chromosomal POX5 gene and the POX4B gene in said C. tropicalis host are disrupted through homologous recombination of each of said POX5 genes and said POX4B gene with a linear DNA fragment comprised of a URA3A selectable marker gene wherein said selectable marker gene is flanked on both ends by DNA sequences having homology to both of said POX5 genes or said POX4B gene respectively.
A transformed C. tropicalis host having an increased copy number of the P450RED gene relative to the wild type strain wherein both copies of the chromosomal POX5 gene and the POX4B gene in said C. tropicalis host are disrupted through homologous recombination of each of said POX5 genes and said POX4B gene with a linear DNA fragment comprised of a URA3A selectable marker gene wherein said selectable marker gene is flanked on both ends by DNA sequences having homology to both of said POX5 genes or said POX4B gene respectively.
A transformed C. tropicalis host having an increased copy number of both the P450ALK and P450RED genes relative to the wild type strain wherein both copies of the chromosomal POX5 gene and the POX4B gene in said C. tropicalis host are disrupted through homologous recombination of each of said POX5 genes and said POX4B gene with a linear DNA fragment comprised of a URA3A selectable marker gene wherein said selectable marker gene is flanked on both ends by DNA sequences having homology to both of said POX5 genes or said POX4B gene respectively.
A transformed C. tropicalis host having an increased copy number of the P450ALK gene relative to the wild type strain wherein one of the chromosomal POX5 genes and the POX4A and POX4B genes in said C. tropicalis host are disrupted through homologous recombination of each of said POX5 genes, said POX4A gene, and said POX4B gene with a linear DNA fragment comprised of a URA3A selectable marker gene wherein said selectable marker gene is flanked on both ends by DNA sequences having homology to said POX5 gene, said POX4A, or said POX4B gene respectively.
A transformed C. tropicalis host having an increased copy number of the P450RED gene relative to the wild type strain wherein one of the chromosomal POX5 genes and the POX4A and POX4B genes in said C. tropicalis host are disrupted through homologous recombination of said POX5 gene, said POX4A, and said POX4B gene each with a linear DNA fragment comprised of a URA3A selectable marker gene wherein said selectable marker gene is flanked on both ends by DNA sequences having homology to said POX5 gene, said POX4A, or said POX4B gene respectively.
A transformed C. tropicalis host having an increased copy number of both the P450ALK and P450RED genes relative to the wild type strain wherein one of the chromosomal POX5 genes and the POX4A and POX4B genes in said C. tropicalis host are disrupted through homologous recombination of said POX5 gene, said POX4A, and said POX4B gene each with a linear DNA fragment comprised of a URA3A selectable marker gene wherein said selectable marker gene is flanked on both ends by DNA sequences having homology to said POX5 gene, said POX4A, or said POX4B gene respectively.
A transformed C. tropicalis host having an increased copy number of the P450ALK gene relative to the wild type strain wherein both copies of the chromosomal POX5 gene and the POX4A and POX4B genes in said C. tropicalis host are disrupted through homologous recombination of each of both of said POX5 genes, said POX4A, and said POX4B gene with a linear DNA fragment comprised of a URA3A selectable marker gene wherein said selectable marker gene is flanked on both ends by DNA sequences having homology to both of said POX5 genes, said POX4A, or said POX4B gene respectively.
A transformed C. tropicalis host having an increased copy number of the P450RED gene relative to the wild type strain wherein both copies of the chromosomal POX5 gene and the and POX4A and POX4B genes in said C. tropicalis host are disrupted through homologous recombination of each of both of said POX5 genes, said POX4A, and said POX4B gene with a linear DNA fragment comprised of a URA3A selectable marker gene wherein said selectable marker gene is flanked on both ends by DNA sequences having homology to both of said POX5 genes, said POX4A, or said POX4B gene respectively.
A transformed C. tropicalis host having an increased copy number of both the P450ALK and P450RED genes relative to the wild type strain wherein both copies of the chromosomal POX5 gene and the POX4A and POX4B genes in said C. tropicalis host are disrupted through homologous recombination of each of both of said POX5 genes, said POX4A, and said POX4B gene with a linear DNA fragment comprised of a URA3A selectable marker gene wherein said selectable marker gene is flanked on both ends by DNA sequences having homology to both of said POX5 genes, said POX4A, or said POX4B gene respectively.
A transformed C. tropicalis host having an increased copy number of the P450ALK gene relative to the wild type strain wherein the chromosomal POX4B and one of the chromosomal POX5 genes in said C. tropicalis host are disrupted through homologous recombination of each of said POX4B and POX5 genes with a linear DNA fragment comprised of a URA3A selectable marker gene wherein said selectable marker gene is flanked on both ends by DNA sequences having homology to said POX4B or one of said POX5 genes respectively.
A transformed C. tropicalis host having an increased copy number of the P450RED gene relative to the wild type strain wherein the chromosomal POX4B and one of the chromosomal POX5 genes in said C. tropicalis host are disrupted through homologous recombination of each of said POX4B and POX5 genes with a linear DNA fragment comprised of a URA3A selectable marker gene wherein said selectable marker gene is flanked on both ends by DNA sequences having homology to said POX4B or one of said POX5 genes respectively.
A transformed C. tropicalis host having an increased copy number of both the P450ALK and P450RED genes relative to the wild type strain wherein the chromosomal POX4B and one of the chromosomal POX5 genes in said C. tropicalis host are disrupted through homologous recombination of each of said POX4B and POX5 genes with a linear DNA fragment comprised of a URA3A selectable marker gene wherein said selectable marker gene is flanked on both ends by DNA sequences having homology to said POX4B or one of said POX5 genes respectively.
A transformed C. tropicalis host having an increased copy number of a P450 gene selected from the group consisting of P450ALK, P450RED and combinations thereof relative to a wild type C. tropicalis strain wherein the level of omega-hydroxylase activity of the transformed C. tropicalis host is at least 0.206 percent lauric acid conversion/.mu.g.
A transformed C. tropicalis host having an increased copy number of a P450RED gene relative to the wild type C. tropicalis strain wherein the omega-hydroxylase activity of the transformed C. tropicalis host is increased relative to the wild type C. tropicalis strain.
Dependencies are inferred from the claim text.