Background: Although CRISPR/Cas enables one-step gene cassette knock-in, assembling targeting vectors containing long homology arms is a laborious process for high-throughput knock-in. We recently developed the CRISPR/Cas-based precise integration into the target chromosome (PITCh) system for a gene cassette knock-in without long homology arms mediated by microhomology-mediated end-joining.
Results: Here, we identified exonuclease 1 (Exo1) as an enhancer for PITCh in human cells. By combining the Exo1 and PITCh-directed donor vectors, we achieved convenient one-step knock-in of gene cassettes and floxed allele both in human cells and mouse zygotes.
Conclusions: Our results provide a technical platform for high-throughput knock-in.
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| http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5126809 | PMC |
| http://dx.doi.org/10.1186/s12864-016-3331-9 | DOI Listing |
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