Simple and Versatile Crispr Based Genome Editing Tool and Its Promise in Large Scale Genome and Cell Engineering Applications

Clustered Regularly Interspaced short palindromic repeat (CRISPR), a bacterial adaptive immune system from Streptococcus pyogenes has been recently engineered and demonstrated to function as a genome editing tool that can rapidly generate engineered cell lines, model organisms and perform large scale gene modifications in a wide variety of hosts. Due to its simple design including Cas9 nuclease and a non-coding guide RNA (gRNA) with target specificity defined by only a short 19-21 base nucleotide region, this system is an attractive tool for large scale genome engineering.  Described here are different CRISPR delivery formats that address both small and large scale genome editing needs in a wide range of host and cell types including stem cells.  Tools and methods tested through this work include a combination of ready to transfect formats that circumvent the need for host specific promoters and reduce the hands on time needed for generating target specific CRISPR system. It also allows the user to edit multiple loci or alleles simultaneously. The simple and easy to design tools described here holds great promise in large scale genome and cell engineering applications.