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US 2010/0075303 A1Published application
Methods for targetted mutagenesis in gram-positive bacteria
AssigneeUT-Battelle, LLC
InventorYunfeng YANG
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Abstract
The present invention provides a method of targeted mutagenesis in Gram-positive bacteria. In particular, the present invention provides a method that effectively integrates a suicide integrative vector into a target gene in the chromosome of a Gram-positive bacterium, resulting in inactivation of the target gene.
Claims21
A method of targeted mutagenesis in a Gram-positive bacterium within the class of Clostridia, comprising:
- providing a suicide integrative vector, wherein said vector is incapable of replication in said bacterium and comprises a targeting nucleotide sequence having a length and sequence identity with a target gene in the chromosome of said bacterium sufficient to permit homologous recombination and integration of said vector into said target gene, wherein integration of said vector into said target gene results in inactivation of said target gene;
- introducing said vector into cells of said bacterium;
- and
- selecting for cells in which said vector has integrated into said target gene.
1 more element
A method of targeted mutagenesis in a Gram-positive bacterium, comprising:
- providing a suicide integrative vector, wherein said vector comprises a γ-origin of replication that is nonfunctional in said bacterium and comprises a targeting nucleotide sequence having a length and sequence identity with a target gene in the chromosome of said bacterium sufficient to permit homologous recombination and integration of said vector into said target gene, wherein integration of said vector into said target gene results in inactivation of said target gene;
- introducing said vector into cells of said bacterium;
- and
- selecting for cells in which said vector has integrated into said target gene.
1 more element
Dependencies are inferred from the claim text.