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Targeted gene insertion into Z chromosome of chicken primordial germ cells for avian sexing model development
Author(s) -
Lee Hong Jo,
Yoon Jong Won,
Jung Kyung Min,
Kim Young Min,
Park Jin Se,
Lee Kyung Youn,
Park Kyung Je,
Hwang Young Sun,
Park Young Hyun,
Rengaraj Deivendran,
Han Jae Yong
Publication year - 2019
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fj.201802671r
Subject(s) - crispr , biology , cas9 , plasmid , genetics , genome editing , gene , sexing , genome , chromosome , microbiology and biotechnology
Clustered regularly interspaced short palindromic repeats (CRISPR) and CRISPR‐associated protein 9 (Cas9) have facilitated the production of genome‐edited animals for use as models. Because of their unique developmental system, avian species offer many advantages as model vertebrates. Here, we report the development of novel chicken models using the CRISPR/Cas9‐mediated nonhomologous end joining repair pathway in chicken primordial germ cells (PGCs). Through the introduction of a donor plasmid containing short guide RNA recognition sequences and CRISPR/Cas9 plasmids into chicken PGCs, exogenous genes of donor plasmids were precisely inserted into target loci, and production of transgenic chickens was accomplished through subsequent transplantation of the Z chromosome–targeted PGCs. Using this method, we successfully accomplished the targeted gene insertion to the chicken sex Z chromosome without detected off‐target effects. The genome‐modified chickens robustly expressed green fluorescent protein from the Z chromosome, which could then be used for easy sex identification during embryogenesis. Our results suggest that this powerful genome‐editing method could be used to develop many chicken models and should significantly expand the application of genome‐modified avians.—Lee, H. J., Yoon, J. W., Jung, K. M., Kim, Y. M., Park, J. S., Lee, K. Y., Park, K. J., Hwang, Y. S., Park, Y. H., Rengaraj, D., Han, J. Y. Targeted gene insertion into Z chromosome of chicken primordial germ cells for avian sexing model development. FASEB J. 33, 8519–8529 (2019). www.fasebj.org