Design of a CRISPR-Cas system to increase resistance of Bacillus subtilis to bacteriophage SPP1
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Clustered regularly interspaced short palindromic repeats (CRISPR) together with CRISPR-associated (cas) genes form an adaptive prokaryotic immune system which provides acquired resistance against viruses and plasmids. Bacillus subtilis presently is the best-characterized laboratory model for Gram-positive bacteria and also widely used for industrial production of enzymes, vitamins and antibiotics. In this study, we show that type II-A CRISPR-Cas system from Streptococcus thermophilus can be transferred into B. subtilis and provides heterologous protection against phage infection. We engineered a heterologous host by cloning S. thermophilus Cas9 and a spacer targeting bacteriophage SPP1 into the chromosome of B. subtilis, which does not harbor its own CRISPR-Cas systems. We found that the heterologous CRISPR-Cas system is functionally active in B. subtilis and provides resistance against bacteriophage SPP1 infection. The high efficiency of the acquired immunity against phage could be useful in generation of biotechnologically important B. subtilis strains with engineered chromosomes.
KeywordsCas9 CRISPR-Cas IMMUNITY Bacillus subtilis Bacteriophage SPP1
We thank Dr. Paulo Tavares (CNRS, Gif-sur-Yvette) and Dr. R. Sapranauskas for kindly provided plasmids.
L Jakutyte-Giraitiene was supported by the postdoctoral fellowship which is being funded by the European Union Structural Funds project “Postdoctoral Fellowship Implementation in Lithuania”. The authors have no conflict of interest to declare.
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