A protocol and webinar detailing use of Edit-R crRNA with electroporation of RNP
Genetic manipulation of primary T cells has the potential for far-reaching implications on human health – everything from novel HIV treatments to personalized cancer therapies. Optimizing the experimental conditions for studies using patient-derived immune cells can be time consuming and subject to variation that are not typical in immortalized cell lines.
A webinar presented by Dr. Judd Hultquist outlined a successful protocol for electroporation of Cas9 ribonuclear protein (RNP) into primary T cells for targeted gene silencing, including high-throughput screening of hundreds of targets. A few key protocol details include:
The experiments presented in the webinar made use of the Edit-R two-part guide RNA platform, utilizing a gene-specific, synthetic crRNA combined with Edit-R tracrRNA, then complexed with Cas9 protein to form the ribonuclear complex (RNP). The ability to receive either custom or predesigned crRNAs in a convenient, customizable 96-well plate format saved a lot of time and handling in generating the RNPs prior to electroporation.
These additional recommendations for consistent and reproducible CRISPR-Cas9 editing in T cells are discussed in the webinar:
Author: Louise Baskin, Senior Product Manager
Guaranteed to edit your target! Algorithm-optimized crRNA for genome-wide coverage of human, mouse, or rat genes. Modifications for nuclease resistance improve DNA-free editing. Simply search for your gene!
Purified Cas9 protein ready to use for DNA-free nuclease expression.
Electroporation protocol for use with Edit-R Cas9 protein NLS and synthetic guide RNA.
Comparing in vitro transcribed and chemically synthesized guide RNA for induction of innate immune response genes.