Application

Functional Genomics/Screening /Target Validation

Biochem/physiol Actions

Utilize the power of CRISPR Whole Genome Lentiviral Pools to knockout and screen every gene in the human genome. The Sigma Whole Human Genome Lentiviral CRISPR Pool is provided as 200 ul in 8 x 25 ul aliquots at a minimum titer of 5x10⁸ viral particles/ml via p24.

Features and Benefits

Use CRISPR nucleases to knockout protein-coding genes to assess their function Efficiently screen the whole human genome (16,000+ genes) at the bench-top without robotics or specialized equipment Increased flexibility consisting of 8 subpools at 23,000 clones per pool (184,000 clones total), from the previous 2 subpools (Gecko v2) Expanded gRNA coverage at 10 gRNAs per gene, from the previous 6 gRNAs per gene (Gecko v2) Numerous built-in enrichment and depletion controls allows researchers to confidently gauge the success of their pooled screening experiments 2 Vector System (pools are gRNA-only, Cas9 sold separately) Ease of optimization: Utilizes same vector system as Gecko v2 allowing for optimization one for both systems Minimize off-targeting: stringent gRNA design and tiling rules

General description

CRISPR (clustered regularly interspaced short palindromic repeat) are repeated sequence of DNA. These are transcribed in a single transcript and later divided into short repeats.

Legal Information

CRISPR Use License AgreementLentiviral and WPRE License Agreements

Other Notes

This product is for R&D use only, not for drug, household, or other uses. Please consult the Material Safety Data Sheet for information regarding hazards and safe handling practices. Though the lentiviral transduction particles produced are replication incompetent, it is recommended that they be treated as Risk Group Level 2 (RGL-2) organisms in laboratory handling. Follow all published RGL-2 guidelines for laboratory handling and waste decontamination.

Preparation Note

Puro Kill Curve and Determining CFU (Colony Formation Unit) per mL. Prior to performing a library-scale screening, two preliminary experiments must be conducted: (1) determine the sensitivity of target cell type to puromycin (kill curve), and (2) determine the functional titer of the lentivirus in your cell type by completing a colony-forming assay (measured in CFU/ml). The calculation of MOI (multiplicity of infection) should be based on the value of CFU. Different cell types vary in transduction efficiency and different lentiviral constructs do not behave identically, so it is critical to optimize your experimental conditions with control lentiviral CRISPR clones (available from Sigma) prior to performing your pooled experiment.