[The cloning of plasmid replicon ori165 from Bacillus thuringiensis subsp. thenebrionis]

An EcoRI DNA fragment containing the plasmid replicon, named ori165, was cloned from the plasmids of B. thuringiensis subsp. tenebrionis strains YBT-1765. The replicon ori165 was the first replicon cloned from this subspecie. Its size is approximately 11 kb. The replicon was minimized to an 8 kb fragment by deletion analysis. Southern hybridization exhibited that ori165 was coming from the plasmid detected with the highest molecular weight. A shuttle vector pBMB6071 based on ori165 was constructed. The stabilities of pBMB6071 in different B. thuringiensis host had great discrepancy. In strain 4Q7, a crystal-negative derivative of B. thuringiensis subsp. isrealensis, the stability was 100%. ori165 was compatible with ori1030 and ori2062 in strain BMB171, a crystal-negative derivative of B. thuringiensis subsp. kurstaki.