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

TLDR

The CRISPR/Cas system enables genome editing by Cas9 guided by a ~20‑nt gRNA, requiring a 5′‑NGG PAM adjacent to the target and genome‑wide specificity to avoid off‑target effects. CRISPRdirect is a web server that selects rational CRISPR/Cas target sites from an input sequence while minimizing off‑target sites through exhaustive genomic searches. The server incorporates genomic sequences from 14 species and performs exhaustive searches to identify target sites with minimal off‑target potential.

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.