PT  - JOURNAL ARTICLE
AU  - Boris V. Skryabin
AU  - Leonid Gubar
AU  - Birte Seeger
AU  - Helena Kaiser
AU  - Anja Stegemann
AU  - Johannes Roth
AU  - Sven G. Meuth
AU  - Hermann Pavenstädt
AU  - Joanna Sherwood
AU  - Thomas Pap
AU  - Roland Wedlich-Söldner
AU  - Cord Sunderkötter
AU  - Yuri B. Schwartz
AU  - Juergen Brosius
AU  - Timofey S. Rozhdestvensky
TI  - Pervasive head-to-tail insertions of DNA templates mask desired CRISPR/Cas9-mediated genome editing events
AID  - 10.1101/570739
DP  - 2019 Jan 01
TA  - bioRxiv
PG  - 570739
4099  - http://biorxiv.org/content/early/2019/03/08/570739.short
4100  - http://biorxiv.org/content/early/2019/03/08/570739.full
AB  - CRISPR/Cas9 mediated homology-directed DNA repair is the method of choice for precise gene editing in a wide range of model organisms, including mouse and human. Broad use by the biomedical community refined the method, making it more efficient and sequence specific. Nevertheless, the rapidly evolving technique still contains pitfalls. During the generation of six different conditional knock-out mouse models, we discovered that frequently (sometimes solely) homology-directed repair and/or non-homologous end-joining mechanisms caused multiple unwanted head-to-tail insertions of donor DNA templates. Disturbingly, conventionally applied PCR analysis—in most cases—failed to identify such multiple integration events, which led to a high rate of falsely claimed precisely edited alleles. We caution that comprehensive analysis of modified alleles is essential, and offer practical solutions to correctly identify precisely edited chromosomes.