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Generation of RUNX 3 knockout pigs using CRISPR /Cas9‐mediated gene targeting
Author(s) -
Kang JT,
Ryu J,
Cho B,
Lee EJ,
Yun YJ,
Ahn S,
Lee J,
Ji DY,
Lee K,
Park KW
Publication year - 2016
Publication title -
reproduction in domestic animals
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.546
H-Index - 66
eISSN - 1439-0531
pISSN - 0936-6768
DOI - 10.1111/rda.12775
Subject(s) - crispr , cas9 , biology , genome editing , gene knockout , somatic cell nuclear transfer , somatic cell , cancer , gene targeting , cancer research , genetically modified organism , knockout mouse , gene , computational biology , genetics , embryogenesis , blastocyst
Contents Pigs are an attractive animal model to study the progression of cancer because of their anatomical and physiological similarities to human. However, the use of pig models for cancer research has been limited by availability of genetically engineered pigs which can recapitulate human cancer progression. Utilizing genome editing technologies such as CRISPR /Cas9 system allows us to generate genetically engineered pigs at a higher efficiency. In this study, specific CRISPR /Cas9 systems were used to target RUNX 3 , a known tumour suppressor gene, to generate a pig model that can induce gastric cancer in human. First, RUNX 3 knockout cell lines carrying genetic modification (monoallelic or biallelic) of RUNX 3 were generated by introducing engineered CRISPR /Cas9 system specific to RUNX 3 into foetal fibroblast cells. Then, the genetically modified foetal fibroblast cells were used as donor cells for somatic cell nuclear transfer, followed by embryo transfer. We successfully obtained four live RUNX 3 knockout piglets from two surrogates. The piglets showed the lack of RUNX 3 protein in their internal organ system. Our results demonstrate that the CRISPR /Cas9 system is effective in inducing mutations on a specific locus of genome and the RUNX 3 knockout pigs can be useful resources for human cancer research and to develop novel cancer therapies.