PLASMID PBRINT - A VECTOR FOR CHROMOSOMAL INSERTION OF CLONED DNA

Plasmid pBRINT is a pBR322 derivative [Bolivar et al., Gene 2 (1977) 95-113; Balbas et al., Gene 50 (1986) 3-40] that allows the insertion and replacement of DNA sequences into the Escherichia coli chromosome by homologous recombination. This method uses the inability of E. coli strain ATCC47002 (JC7623) to replicate covalently closed circular (ccc) pBR322-derived plasmids, and the convenience of XGal+IPTG screening for recombinants. The vector also contains suitable selection markers (Ap and Cm), as well as a multiple cloning site (MCS) derived from the pUC vectors [Yanisch-Perron et al., Gene 33 (1985) 103-119] to facilitate cloning. A simple PCR scheme was developed to scan for DNA insertions into the bacterial chromosome. Once introduced into the chromosome, the inserted DNA sequences can be transferred to other strains by bacteriophage P1-mediated transduction.