USER friendly cloning coupled with chitin-based natural transformation enables rapid mutagenesis of Vibrio vulnificus.

ABSTRACT: Vibrio vulnificus is a bacterial contaminant of shellfish and causes highly lethal sepsis and destructive wound infections. A definitive identification of virulence factors using the molecular version of Koch's postulates has been hindered because of difficulties in performing molecular genetic analysis of this opportunistic pathogen. For example, conjugation is required to introduce plasmid DNA, and allelic exchange suicide vectors that rely on sucrose sensitivity for counterselection are not efficient. We therefore incorporated USER friendly cloning techniques into pCVD442-based allelic exchange suicide vectors and other expression vectors to enable the rapid and efficient capture of PCR amplicons. Upstream and downstream DNA sequences flanking genes targeted for deletion were cloned tog

SUBMITTER: Gulig PA 

PROVIDER: S-EPMC2725515 | biostudies-literature | 2009 Aug

REPOSITORIES: biostudies-literature