| First Author | Gennequin B | Year | 2013 |
| Journal | Biochem Biophys Res Commun | Volume | 441 |
| Issue | 4 | Pages | 815-9 |
| PubMed ID | 24211574 | Mgi Jnum | J:211501 |
| Mgi Id | MGI:5575591 | Doi | 10.1016/j.bbrc.2013.10.138 |
| Citation | Gennequin B, et al. (2013) CRISPR/Cas-induced double-strand breaks boost the frequency of gene replacements for humanizing the mouse Cnr2 gene. Biochem Biophys Res Commun 441(4):815-9 |
| abstractText | The CRISPR/Cas technology has been successfully used to stimulate the integration of small DNA sequences in a target locus to produce gene mutations. However, many applications require homologous recombination using large gene-targeting constructs. Here we address the potential of CRISPR/Cas-mediated double-strand breaks to enhance the genetic engineering of large target sequences using a construct for "humanizing" the mouse Cnr2 gene locus. We designed a small-guide RNA that directs the induction of double strand breaks by Cas9 in the Cnr2 coding exon. By co-transfection of the CRISPR/Cas system with the 10 kb targeting construct we were able to boost the recombination frequency more than 200-fold from 0.27% to 67%. This simple technology can thus be used for the homologous integration of large gene fragments and should greatly enhance our ability to generate any kind of genetically altered mouse models. |
