Bacillus subtilis has become a widely used microbial cell factory for the production of recombinant proteins, especially those associated with foods and food processing. Recent advances in genetic manipulation and proteomic analysis have been used to greatly improve protein production in B. subtilis. This review begins with a discussion of genome-editing technologies and application of the CRISPR–Cas9 system to B. subtilis. A summary of the characteristics of crucial legacy strains is followed by suggestions regarding the choice of origin strain for genetic manipulation. Finally, the review analyzes the genes and operons of B. subtilis that are important for the production of secretory proteins and provides suggestions and examples of how they can be altered to improve protein production. This review is intended to promote the engineering of this valuable microbial cell factory for better recombinant protein production.

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