CRISPRi-FGP: web-based genome-scale CRISPRi sgRNA design and validation tool in prokaryotes

Pooled CRISPRi with high-throughput sequencing is emerging as a novel functional genomic study method in prokaryotes. Due to the diversity of sgRNA activity and off-target effect, it is crucial to have a robust sgRNA library designing tool. Here we propose a web tool named CRISPRi-FGP (http://www.thu-big.net/sgRNA_design/), to assist sgRNA library design in prokaryotes. CRISPRi-FGP applies a build-in database containing 1,300 common prokaryotes as standard designing source, while also accepts customized designing. It employs a scoring metrics based on experiment data, together with specially trimed algorithm for off-target selection both in high quality and speed. CRISPRi-FGP exhibited high accuracy and robustness on sgRNA library designing in multiple prokaryotes. Finally, an experiment comparing CRISPRi-FGP and existing tool was conducted and showed CRISPRi-FGP has improved ability in recognizing off-target sites. All these results showed CRISPRi-FGP with the potential to be a precise and robust tool for high-quality sgRNA library design in prokaryotes.