論文

国際誌
2021年1月

Introduction of a point mutation in the KRAS gene of in vitro fertilized porcine zygotes via electroporation of the CRISPR/Cas9 system with single-stranded oligodeoxynucleotides.

Animal science journal = Nihon chikusan Gakkaiho
  • Manita Wittayarat
  • ,
  • Maki Hirata
  • ,
  • Zhao Namula
  • ,
  • Yoko Sato
  • ,
  • Nhien T Nguyen
  • ,
  • Quynh A Le
  • ,
  • Qingyi Lin
  • ,
  • Koki Takebayashi
  • ,
  • Fuminori Tanihara
  • ,
  • Takeshige Otoi

92
1
開始ページ
e13534
終了ページ
記述言語
英語
掲載種別
研究論文(学術雑誌)
DOI
10.1111/asj.13534

This study aimed to investigate the efficiency of KRAS gene editing via CRISPR/Cas9 delivery by electroporation and analyzed the effects of the non-homologous end-joining pathway inhibitor Scr7 and single-stranded oligodeoxynucleotide (ssODN) homology arm length on introducing a point mutation in KRAS. Various concentrations (0-2 µM) of Scr7 were evaluated; all concentrations of Scr7 including 0 µM resulted in the generation of blastocysts with a point mutation and the wild-type sequence or indels. No significant differences in the blastocyst formation rates of electroporated zygotes were observed among ssODN homology arm lengths, irrespective of the gRNA (gRNA1 and gRNA2). The proportion of blastocysts carrying a point mutation with or without the wild-type sequence and indels was significantly higher in the ssODN20 group (i.e., the group with a ssODN homology arm of 20 bp) than in the ssODN60 group (gRNA1: 25.7% vs. 5.4% and gRNA2: 45.5% vs. 5.9%, p < .05). In conclusion, the CRISPR/Cas9 delivery with ssODN via electroporation is feasible for the generation of point mutations in porcine embryos. Further studies are required to improve the efficiency and accuracy of the homology-directed repair.

リンク情報
DOI
https://doi.org/10.1111/asj.13534
PubMed
https://www.ncbi.nlm.nih.gov/pubmed/33638256
ID情報
  • DOI : 10.1111/asj.13534
  • PubMed ID : 33638256

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