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Production of non-mosaic genome edited porcine embryos by injection of CRISPR/Cas9 into germinal vesicle oocytes

摘要Genetically modified pigs represent a great promise for generating models of human diseases and producing new breeds.Generation of genetically edited pigs using somatic cell nuclear transfer (SCNT) or zygote cytoplasmic microinjection is a tedious process due to the low developmental rate or mosaicism of the founder (F0).Herein,we developed a method termed germinal vesicle oocyte gene editing (GVGE) to produce non-mosaic porcine embryos by editing maternal alleles during the GV to MII transition.Injection of Cas9 mRNA and X-linked Dmd gene-specific gRNA into GV oocytes did not affect their developmental potential.The MII oocytes edited during in vitro maturation (IVM) could develop into blastocysts after parthenogenetic activation (PA) or in vitro fertilization (IVF).Genotyping results indicated that the maternal gene X-linked Dmd could be efficiently edited during oocyte maturation.Up to 81.3% of the edited IVF embryos were non-mosaic Dmd gene mutant embryos.In conclusion,GVGE might be a valuable method for the generation of non-mosaic maternal allele edited F0 embryos in a short simple step.

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作者 Xiaohu Su [1] Wei Chen [2] Qingqing Cai [2] Puping Liang [1] Yaosheng Chen [2] Peiqing Cong [2] Junjiu Huang [3] 学术成果认领
作者单位 Key Laboratory of Reproductive Medicine of Guangdong Province, First Affiliated Hospital and School of Life Sciences, Sun Yat-sen University, Guangzhou,510275, China;MOE Key Laboratory of Gene Function and Regulation, State Key Laboratory of Biocontrol, School of Life Sciences, Sun Yat-sen University, Guangzhou,510275, China [1] MOE Key Laboratory of Gene Function and Regulation, State Key Laboratory of Biocontrol, School of Life Sciences, Sun Yat-sen University, Guangzhou,510275, China [2] Key Laboratory of Reproductive Medicine of Guangdong Province, First Affiliated Hospital and School of Life Sciences, Sun Yat-sen University, Guangzhou,510275, China;MOE Key Laboratory of Gene Function and Regulation, State Key Laboratory of Biocontrol, School of Life Sciences, Sun Yat-sen University, Guangzhou,510275, China;Key Laboratory of Reproductive Medicine of Guangdong Province, Third Affiliated Hospital of Guangzhou Medical University, Guangzhou, 510150, China [3]
栏目名称 Original Research
发布时间 2020-11-23
基金项目
This work was supported by the National Key R&D Program of China the National Natural Science Foundation of China the National Transgenic Major Program the Natural Science Foundation of Guangdong Province the Key R&D Program of Guangdong Province and the Guangzhou Science and Technology Project
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