Cloning of the neutral protease gene of Bacillus subtilis and the use of the cloned gene to create an in vitro-derived deletion mutation

The neutral protease gene of Bacillus subtilis has been cloned, and its nucleotide sequence has been determined. The cloned gene was used to create an in vitro-derived deletion mutation, which was used to replace the wild-type copy of the gene. This deletion, in combination with a deletion of the alkaline protease gene, completely abolished protease production. The loss of the proteases had no detectable effect on growth, morphology, or sporulation.