CRISPR RNA-dependent binding and cleavage of endogenous RNAs by the Campylobacter jejuni Cas9

This study investigates the RNA targets and cleavage sites of endogenous Cas9 in the food-borne pathogen Campylobacter jejuni. Direct RNA binding targets of Cas9 in C. jejuni strain NCTC11168 were determined using RIP-seq. The Cleavage sites were then predicted in the RNA targets by comparing total transcriptome data from WT and deletion (cas9, crRNA3, tracrRNA, CRISPR-tracrRNA) strains. PAMs for the CjeCas9 were enriched using the PAM-SCANR platform, which operates through a GFP reporter gene. Upon GFP (and thus functional PAM) enrichment, fluorescing cells were isolated using FACS and prepared plasmid DNA was amplified and prepared for sequencing. Identification of Cas9 binding and cleavage sites in C. jejuni strain NCTC11168. Four total files are included for the PAM-SCANR dataset. Raw reads are available for both biological replicates, both pre- and post-FACS. After extracting the variable PAM region from the reads, enrichment was calculated by comparing the pre-FACS PAM counts to the post-FACS counts. These calculations are presented in the included processed data file.