FIG 2.

The CRISPR1-Cas system of S. mutans UA159 provides immunity against plasmid transformation. (A) Schematic representation of cloning vector pCG1 used for the construction of plasmids for the transformation interference assay. Plasmids for interference assays were produced by inserting a protospacer and 10 nt on both sides of the protospacers into pCG1 plasmid. (B) pCG1 constructs containing potential targets for different S. mutans UA159 spacers (spacers 2, 3, and 6 within CRISPR1 array and spacer 1 within CRISPR2 array). pCG1derivatives in the presence (C) or absence (D) of flank sequences were tested by natural transformation assays using the wild-type S. mutans UA159, ΔC1K, ΔC2E, and ΔC1SC2E strains. The transformation frequency was calculated as the transformant CFU divided by the total number of viable cells. The results shown are representative of at least two independent experiments. ***, constructs showing targeting phenotype.