A Simple Method for Introducing Marker-Free Deletions in the Bacillus subtilis Genome

Abstract

A genetic tool for introducing marker-free deletions is essential for multiple manipulations of genomes. We have developed a simple and efficient method for creating marker-free deletion mutants of Bacillus subtilis through transformation with recombinant PCR products, using the Escherichia coli mazF gene encoding an endoribonuclease that cleaves free mRNAs as a counterselection tool.