Non-revertible beta-oxidation blocked candida tropicalis
AssigneeCognis Corporation
InventorDavid L. Craft
Life of this patent
Estimating the term…
Abstract
Genetically modified strains of C. tropicalis, which will not revert to wild-type activity at the POX 4 and/or POX 5 locus, are disclosed. The strains are β-oxidation blocked and have been transformed through homologous recombination with a construct which deletes a portion of the POX 4 and/or POX 5 gene. The modified strains may be used to increase yields of dicarboxylic acids produced in host cells of the strains. Methods for blocking the β-oxidation pathway in a C. tropicalis host cell are also provided.
Claims43
A Candida tropicalis cell having a disrupted chromosomal POX 4 gene wherein a portion of said POX 4 gene has been deleted through homologous recombination with a DNA fragment comprising a selectable marker gene, wherein said selectable marker gene is flanked on both ends by DNA sequences having homology to non-contiguous portions of said POX 4 gene and wherein the presence of the deleted portion of the POX 4 gene prevents recombination to a functional POX 4 gene.
A Candida tropicalis cell having a disrupted chromosomal POX 5 gene wherein a portion of said POX 5 gene has been deleted through homologous recombination with a DNA fragment comprising a selectable marker gene, wherein said selectable marker gene is flanked on both ends by DNA sequences having homology to non-contiguous portions of said POX 5 gene and wherein the presence of the deleted portion of the POX 5 gene prevents recombination to a functional POX 4 gene.
A Candida tropicalis cell having disrupted chromosomal POX 4 and POX 5 genes wherein a portion of both of said POX 4 and POX 5 genes has been deleted through homologous recombination with a DNA fragment comprising a selectable marker gene wherein said selectable marker gene is flanked on both ends by DNA sequences having homology to non-contiguous portions of both of said POX 4 and POX 5 genes and wherein the presence of the deleted portions of both of the POX 4 and POX 5 genes prevent recombination to functional POX 4 and POX 5 genes.
A process for completely blocking the β-oxidation pathway in a Candida tropicalis host cell which comprises disrupting through homologous recombination a portion of a POX 4 gene with a DNA fragment comprising a selectable marker gene wherein said selectable marker gene is flanked on both ends by DNA sequences having homology to non-contiguous portions of said POX 4 gene and wherein the presence of the deleted portion of the POX 4 gene prevents recombination to a functional POX 4 gene.
A process for completely blocking the β-oxidation pathway in a Candida tropicalis host cell which comprises disrupting through homologous recombination a portion of a POX5 gene with a DNA fragment comprising a selectable marker gene wherein said selectable marker gene is flanked on both ends by DNA sequences having homology to non-contiguous parts of said POX 5 gene and wherein the presence of the deleted portion of the POX 5 gene prevents recombination to a functional POX 5 gene.
A process for completely blocking the β-oxidation pathway in a Candida tropicalis host cell which comprises disrupting through homologous recombination a portion of both of POX 4 and POX 5 genes with a DNA fragment comprised of a selectable marker gene wherein said selectable marker gene is flanked on both ends by DNA sequences having homology to non-contiguous portions of both of said POX 4 and POX 5 genes and wherein the deleted portions of both of the POX 4 and POX 5 genes prevent recombination to functional POX 4 and POX 5 genes.
Dependencies are inferred from the claim text.