Title

CRISPRtools: a flexible computational platform for performing CRISPR/Cas9 experiments in the mouse.

Document Type

Article

Publication Date

8-2017

JAX Source

Mamm Genome 2017 Aug; 28(7-8):283-290

PMID

28280930

DOI

https://doi.org/10.1007/s00335-017-9681-z

Grant

OD011185, OD023222, OD010972

Abstract

Genome editing using the CRISPR/Cas9 RNA-guided endonuclease system has rapidly become a driving force for discovery in modern biomedical research. This simple yet elegant system has been widely used to generate both loss-of-function alleles and precision knock-in mutations using single-stranded donor oligonucleotides. Our CRISPRtools platform supports both of these applications in order to facilitate the use of CRISPR/Cas9. While there are several tools that facilitate CRISPR/Cas9 design and screen for potential off-target sites, the process is typically performed sequentially on single genes, limiting scalability for large-scale programs. Here, the design principle underlying gene ablation is based upon using paired guides flanking a critical region/exon of interest to create deletions. Guide pairs are rank ordered based upon published efficiency scores and off-target analyses, and reported in a concise format for downstream implementation. The exon deletion strategy simplifies characterization of founder animals and is the strategy employed for the majority of knockouts in the mouse. In proof-of-principle experiments, the effectiveness of this approach is demonstrated using microinjection and electroporation to introduce CRISPR/Cas9 components into mouse zygotes to delete critical exons. Mamm Genome 2017 Aug; 28(7-8):283-290.