徳島大学 教育・研究者情報データベース(EDB)

1. (英) Lin Qingyi

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2. (英) Le Quynh Anh

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3. (英) Takebayashi Koki

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4. ソンキテイデイロ チヨンマナート

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5. (英) Wittayarat Manita

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6. 平田 真樹([徳島大学.バイオイノベーション研究所])

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7. (英) Tanihara Fuminori

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8. 音井 威重([徳島大学.バイオイノベーション研究所])

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(英) Timing and duration of lipofection-mediated CRISPR/Cas9 delivery into porcine zygotes affect gene-editing events.

(英) Lipofection-mediated introduction of the CRISPR/Cas9 system in porcine zygotes provides a simple method for gene editing, without requiring micromanipulation. However, the gene editing efficiency is inadequate. The aim of this study was to improve the lipofection-mediated gene editing efficiency by optimizing the timing and duration of lipofection.

(日) Zona pellucida (ZP)-free zygotes collected at 5, 10, and 15 h from the start of in vitro fertilization (IVF) were incubated with lipofection reagent, guide RNA (gRNA) targeting GGTA1, and Cas9 for 5 h. Lipofection of zygotes collected at 10 and 15 h from the start of IVF yielded mutant blastocysts. Next, ZP-free zygotes collected at 10 h from the start of IVF were incubated with lipofection reagent, gRNA, and Cas9 for 2.5, 5, 10, or 20 h. The blastocyst formation rate of zygotes treated for 20 h was significantly lower (p < 0.05) than those of the other groups, and no mutant blastocysts were obtained. Moreover, the mutation rates of the resulting blastocysts decreased as the incubation time increased. In conclusion, a lipofection-mediated gene editing system using the CRISPR/Cas9 system in ZP-zygotes is feasible; however, further improvements in the gene editing efficiency are required.

1. (英) Animals
2. (英) Blastocyst
3. (英) CRISPR-Cas Systems
4. (英) Gene Editing
5. (英) RNA, Guide
6. (英) Swine
7. (英) Zygote

1. (英) PublicationType: Journal Article