The Guide-it CRISPR Genome-Wide sgRNA Library System makes it easy to perform lentiviral-based, genome-wide phenotypic knockout screens using CRISPR/Cas9 in human cell lines. Each screen contains both the genome-wide sgRNA library and a Cas9 expression system in our easy-to-use Lenti-X Single Shots format to produce cell lines stably expressing both Cas9 and the sgRNA library. The library is based on the Brunello library (Doench et al. 2016; Doench, 2018), and contains >76,000 guide RNAs (sgRNAs) with four highly active guides per gene, targeting about 19,000 genes as well as non-targeting sgRNA controls.
After performing a screen, use next-generation sequencing (NGS) analysis with the Guide-it CRISPR Genome-Wide sgRNA Library NGS Analysis Kit, which includes the necessary primers and kits to prepare an NGS sample from cells transduced with our sgRNA library.
This library has been carefully manufactured to ensure proper sgRNA representation. We have measured sgRNA representation at every step in the process: library synthesis, amplification, viral production, and target-cell transduction. This kit contains the validated library in a dry-down lentiviral format and Lenti-X 293T cells. To produce viral particles, simply add water to the lyophilized library mix and then add it to the 293T cells in culture. The sgRNA content is verified in every lot via NGS analysis to contain >90% of all the stated sgRNAs within a 10-fold distribution range. We have also modified the sgRNA scaffold design in the pLVXS-sgRNA-mCherry-hyg Vector to improve the Cas9-sgRNA interaction, ensuring high editing efficiency. Representation of the lentiviral sgRNA library is maintained by utilizing our Lenti-X Packaging Single Shots, a validated, high-titer packaging formulation.
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