CRISPRdirect: software for designing CRISPR/Cas guide RNA with reduced off-target sites

Abstract

CRISPRdirect is a simple and functional web server for selecting rational CRISPR/Cas targets from an input sequence. The CRISPR/Cas system is a promising technique for genome engineering which allows target-specific cleavage of genomic DNA guided by Cas9 nuclease in complex with a guide RNA (gRNA), that complementarily binds to a ∼ 20 nt targeted sequence. The target sequence requirements are twofold. First, the 5'-NGG protospacer adjacent motif (PAM) sequence must be located adjacent to the target sequence. Second, the target sequence should be specific within the entire genome in order to avoid off-target editing. CRISPRdirect enables users to easily select rational target sequences with minimized off-target sites by performing exhaustive searches against genomic sequences. The server currently incorporates the genomic sequences of human, mouse, rat, marmoset, pig, chicken, frog, zebrafish, Ciona, fruit fly, silkworm, Caenorhabditis elegans, Arabidopsis, rice, Sorghum and budding yeast.

Availability: Freely available at http://crispr.dbcls.jp/.

Supplementary information: Supplementary data are available at Bioinformatics online.

Fig. 1.

Screenshot from the CRISPRdirect web server. (A) Top page. The server accepts either an accession number or a nucleotide sequence as input. (B) Typical output of CRISPRdirect. A list of CRISPR/Cas target candidates is displayed. (C) A graphical view of target sites demonstrates the position and orientation of each site. (D) The results can be exported as tab-delimited text or in JSON format. (E) Detailed list of potential off-target sites which visualizes the positions of mismatches and gaps