Construction and characterization of the host component of the Bacillus subtilis HV2 cloning system.

Abstract

A strain of Bacillus subtilis has been developed and tested for use as the host component of the B. subtilis HV2 system. The strain was constructed incorporating the following mutations: thyA, thyB, uvr-1, dal, spoOA delta 677, citD delta 20 (uvrC), strd. The strain is unable to grow in the absence of thymine, D-alanine, or streptomycin; unable to sporulate; and sensitive to UV radiation. Survival of the attenuated strain was severely restricted in nonpermissive environments, and the organism suffered a five-log decrease in viability within 24 h when dried at room temperature. The strain could be transformed with plasmid DNA using the protoplast transformation system and was transducible with PBS1. The ability of the strain to serve as host for the temperate phages phi 3T and rho 11 and for the plasmid vectors pUB110 and pBD64 was unimpaired.