Cloning of sporulation gene spoIVC in Bacillus subtilis

Abstract

Sporulation gene spoIVC of Bacillus subtilis was cloned by the prophage transformation method in temperate phage phi 105. The specialized transducing phage, phi 105 spoIVC-1, restored the sporulation of the asporogenous mutant of B. subtilis strain 1S47 (spoIVC133). Transformation experiments showed that the spoIVC gene resides on a 7.3 kb HindIII restriction fragment. Subsequent analysis of the 7.3 kb HindIII fragment with restriction endonuclease EcoRI showed that the spoIVC gene resides on a 3.6 kb EcoRI fragment within the 7.3 kb fragment. The 3.6 kb fragment was recloned into the unique EcoRI site of plasmid pUB110 and deletion derivatives having a deletion within the 3.6 kb insert were constructed. The plasmid carrying the entire spoIVC gene restored the sporulation of strain HU1214 (spoIVC133, recE4) at a frequency of 10(7) spores/ml, and reduced the sporulation of strain HU1018 (spo+, recE4) to 10(7) spores/ml.