Functional characterization of cas4 of a type I-D CRISPR-Cas system in vivo.

We introduced the CRISPR adaptation genes cas1, cas2 and cas4 from the Type I-D CRISPR-Cas system of Synechocystis sp. 6803 into Escherichia coli and analyzed the CRISPR spacers that were acquired in either the presence or absence of cas4 by high-throughput sequencing. We observed that cas4 is strictly required for the selection of targets with protospacer adjacent motifs (PAMs) conferring I-D CRISPR interference in the native host Synechocystis.