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phytoCRISP-Ex is a rational and flexible tool for finding Cas9 target sites with low/no off-targeting potential. Given a DNA query (eg. Gene) sequence, the pipeline first fetches all the possible regions of length 23, structured as [5p- G(19bps)NGG -3p]. Further these regions are evaluated and filtered to show low/no off-target activity across the whole genome.

There are two levels of filters, passing both designates the region a potential sgRNA for Cas9 activity. The first level rejects the target and tag it "FAIL" if it has more than 2 matches with the off-targets in the genome and second filter, which runs in parallel, rejects the target if the seed region (last 15 bases including the PAM sequence) has 100% match with the off-targets anywhere in the genome. The pipeline reports only the instances which are accepted by both the filters and are designated as potential Cas9 targets.

phytoCRISP-Ex pipeline is mounted with the index of 13 Protist genomes and is accessible via web-server. An off-line standalone software is also developed which gives liberty to its users to use it for any genome, assembled or un-assembled.

We always work on improvements in our programs, to make them better, smarter and user friendly. Thus, feel free to post valuable suggestions, bugs or troubleshooting queries via email at rastogi@biologie.ens.fr

Institut de Biologie de l'ENS (IBENS)