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

Xiaohu Su^{a, b, #},
Wei Chen^{b, #},
Qingqing Cai^b,
Puping Liang^{a, b},
Yaosheng Chen^b,
Peiqing Cong^{b
, ,},
Junjiu Huang^{a, b, c
, ,}

a.
Key Laboratory of Reproductive Medicine of Guangdong Province, First Affiliated Hospital and School of Life Sciences, Sun Yat-sen University, Guangzhou, 510275, China
b.
MOE Key Laboratory of Gene Function and Regulation, State Key Laboratory of Biocontrol, School of Life Sciences, Sun Yat-sen University, Guangzhou, 510275, China
c.
Key Laboratory of Reproductive Medicine of Guangdong Province, Third Affiliated Hospital of Guangzhou Medical University, Guangzhou, 510150, China

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Corresponding author: E-mail address: congpq@mail.sysu.edu.cn (Peiqing Cong); E-mail address: hjunjiu@mail.sysu.edu.cn (Junjiu Huang)

These authors contributed equally to this work.

摘要

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Abstract: 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.
- Porcine /
- Germinal vesicle (GV) /
- CRISPR/Cas9
These authors contributed equally to this work.
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doi: 10.1016/j.jgg.2019.07.002

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

Corresponding author: E-mail address: congpq@mail.sysu.edu.cn (Peiqing Cong); E-mail address: hjunjiu@mail.sysu.edu.cn (Junjiu Huang)

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doi: 10.1016/j.jgg.2019.07.002

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

Corresponding author: E-mail address: congpq@mail.sysu.edu.cn (Peiqing Cong); E-mail address: hjunjiu@mail.sysu.edu.cn (Junjiu Huang)

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