Which genes are targeted using your library? Where can I find a list of the targeted genes?

Guide RNAs in our library target the entire human genome of 19,114 genes (including essential genes). There are four guides per gene, which, including controls amounts to 76,612 different sgRNA vector constructs. A complete list of all sgRNAs in our library and their representation data can be requested at takarabio.com/genomewidelibrary. http://www.takarabio.com/genomewidelibraryGene editing [...]

What is the design of the vector backbones used in your library?

Both our sgRNA library and the Cas9 sequence are cloned into a lentiviral vector backbone consisting of a 3' self-inactivating LTR and sequence elements (e.g., cPPT/CTS and WPRE) to enhance titer and transduction efficiency. Cas9 (from S.pyogenes) is expressed from the human EF1-alpha promoter, and stable integrants can be selected using puromycin. The sgRNA library is expressed from a human U6 [...]

What are the advantages of using an sgRNA library in Takara Bio’s Lenti-X Single Shots format vs. starting from plasmid or lentiviral particles?

Researchers conducting pooled, lentiviral CRISPR knockout screens often start either with an sgRNA library in plasmid form or premade lentiviral particles. Starting from plasmid involves a much lower upfront cost, but the process of amplifying the library, verifying sgRNA representation, and producing lentivirus require familiarity with a wide array of techniques and often proves to be [...]

The Guide-it library is supplied in a lyophilized Lenti-X Single Shots format. What does that mean?

The Lenti-X packaging single shots format provides a straightforward and consistent method to package high-titer lentivirus. The lyophilized pellet (see image below) contains our fourth-generation lentiviral packaging system, a premixed formulation of five Lenti-X packaging plasmids and the library (>76,000 sgRNA lentiviral plasmids) all combined in a single tube at preoptimized ratios. No [...]

What are the kit components of the Guide-it CRISPR Genome-Wide sgRNA Library System?

There are five components to the library system as described below: 632646 Guide-it CRISPR Genome-Wide sgRNA Library System, 5 Screens -Guide-it Genome-Wide sgRNA Library Transfection Mix, 2 x 5 vials -Guide-it Cas9 Lentiviral Transfection Mix, 5 vials -pLVXS-sgRNA-mCherry-hyg Vector, 5ug -Lenti-X 293T Cell Line, 2 x 106 cells -Lenti-X GoStix Plus (Sample), 3 tests 1. The Guide-it Genome-Wide [...]