Project description: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.
Project description:Off-target (OT) analysis of different guide RNAs targeting PKLR gene. GUIDE-Seq analyses were done in HEK293 cells stably expressing Cas9 transfected with gRNA complexes, generated using Alt-R® CRISPR-Cas9 crRNA XT and Alt-R®CRISPR-Cas9 tracrRNA, and transfected with a dsODN tag
Project description:In this study the RNA targetome of the endonuclease Cas9, the trans-activating crRNA (tracrRNA) and a CRISPR-associated small RNA (scaRNA/NMnc0040) was determined in Neisseria meninigitidis serogroup C strain 8013 by RNA-seq.
