Abstract: Provided are a system for constructing a genome-wide sgRNA library and a use thereof. The system includes an input module, an sgRNA design module and an sgRNA filtering module. By constructing three modules in the system, optimizing details and processes in the modules, and adopting multiple design criteria and screening principles, the genome-wide sgRNA library is finally constructed. The system and method are concise and efficient, and the obtained library has a high quality and good activity, and is convenient for applications in gene editing researches.