TYPE III-A CRISPR/CAS SYSTEMS AND COMPARISON OF CAS1, CAS2, AND CAS10 PROTEINS OF LACTOBACILLI |

In lactic acid bacteria, type I and type II CRISPR systems are the most common, however type III is found in a few taxa. Lactobacilli with a type III-A CRISPR system were identified and their Cas10 signature nucleases, Cas1, Cas2 proteins, and repeat sequences were compared in this study. All lactobacilli in 39 strains from 9 species possessed the Type III-A CRISPR system. Among all species, Ligilactobacillus salivarius strains possessed the most type III-A systems. The most common repeat sequence was “agtaagaacatatctccgaatataggagacgaaac,” and the secondary structure was predicted using the RNAfold analysis tool. The phylogenetic tree and amino acid sequence alignment of Cas1, Cas2, and Cas10 revealed that the strains tended to break into species-specific branches. Between cas6 and CRISPR array, L. ruminis ATCC 27782 and ATCC 27780 possessed IS66 family insertion sequence element accessory protein TnpB, as well as DEDDh genes. Type III-A CRISPR systems' endogenous Cas10 enzyme has recently been used in genetic engineering experiments, and reprogrammed type III-A systems have been tested for endogenous genome editing. As a result, this work retraced lactobacilli type III-A CRISPR systems to assess clustering of repeat sequences, Cas1, Cas2, and Cas10 proteins among strains, as well as to identify lactobacilli as a potential for future reprogramming endogenous genome editing.

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