PT  - JOURNAL ARTICLE
AU  - Wesley A. Wierson
AU  - Jordan M. Welker
AU  - Maira P. Almeida
AU  - Carla M. Mann
AU  - Dennis A. Webster
AU  - Melanie E. Torrie
AU  - Trevor J. Weiss
AU  - Macy K. Vollbrecht
AU  - Merrina Lan
AU  - Kenna C. McKeighan
AU  - Zhitao Ming
AU  - Alec Wehmeier
AU  - Christopher S. Mikelson
AU  - Jeffrey A. Haltom
AU  - Kristen M. Kwan
AU  - Chi-Bin Chien
AU  - Darius Balciunas
AU  - Stephen C. Ekker
AU  - Karl J. Clark
AU  - Beau R. Webber
AU  - Branden Moriarity
AU  - Staci L. Solin
AU  - Daniel F. Carlson
AU  - Drena L. Dobbs
AU  - Maura McGrail
AU  - Jeffrey J. Essner
TI  - GeneWeld: a method for efficient targeted integration directed by short homology
AID  - 10.1101/431627
DP  - 2018 Jan 01
TA  - bioRxiv
PG  - 431627
4099  - http://biorxiv.org/content/early/2018/10/24/431627.short
4100  - http://biorxiv.org/content/early/2018/10/24/431627.full
AB  - Choices for genome engineering and integration involve high efficiency with little or no target specificity or high specificity with low activity. Here, we describe a targeted integration strategy, called GeneWeld, and a vector series for gene tagging, pGTag (plasmids for Gene Tagging), which promote highly efficient and precise targeted integration in zebrafish embryos, pig fibroblasts, and human cells utilizing the CRISPR/Cas9 system. Our work demonstrates that in vivo targeting of a genomic locus of interest with CRISPR/Cas9 and a donor vector containing as little as 24 to 48 base pairs of homology directs precise and efficient knock-in when the homology arms are exposed with a double strand break in vivo. Given our results targeting multiple loci in different species, we expect the accompanying protocols, vectors, and web interface for homology arm design to help streamline gene targeting and applications in CRISPR compatible systems.In Brief Wierson et al. describe a targeted integration strategy, called GeneWeld, and a vector series for gene tagging, pGTag, which promote highly efficient and precise targeted integration in zebrafish, pig fibroblasts, and human cells. This approach establishes an effective genome engineering solution that is suitable for creating knock-in mutations for functional genomics and gene therapy applications. The authors describe high rates of germline transmission (50%) for targeted knock-ins at eight different zebrafish loci and efficient integration at safe harbor loci in porcine and human cells.