Efficient generation of Knock-in/Knock-out marmoset embryo via CRISPR/Cas9 gene editing

Wakako Kumita, Kenya Sato, Yasuhiro Suzuki, Yoko Kurotaki, Takeshi Harada, Yang Zhou, Noriyuki Kishi, Kengo Sato, Atsu Aiba, Yasubumi Sakakibara, Guoping Feng, Hideyuki Okano, Erika Sasaki

Research output: Contribution to journalArticle

Abstract

Genetically modified nonhuman primates (NHP) are useful models for biomedical research. Gene editing technologies have enabled production of target-gene knock-out (KO) NHP models. Target-gene-KO/knock-in (KI) efficiency of CRISPR/Cas9 has not been extensively investigated in marmosets. In this study, optimum conditions for target gene modification efficacies of CRISPR/mRNA and CRISPR/nuclease in marmoset embryos were examined. CRISPR/nuclease was more effective than CRISPR/mRNA in avoiding mosaic genetic alteration. Furthermore, optimal conditions to generate KI marmoset embryos were investigated using CRISPR/Cas9 and 2 different lengths (36 nt and 100 nt) each of a sense or anti-sense single-strand oligonucleotide (ssODN). KIs were observed when CRISPR/nuclease and 36 nt sense or anti-sense ssODNs were injected into embryos. All embryos exhibited mosaic mutations with KI and KO, or imprecise KI, of c-kit. Although further improvement of KI strategies is required, these results indicated that CRISPR/Cas9 may be utilized to produce KO/KI marmosets via gene editing.

Original languageEnglish
Article number12719
JournalScientific reports
Volume9
Issue number1
DOIs
Publication statusPublished - 2019 Dec 1

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Clustered Regularly Interspaced Short Palindromic Repeats
Callithrix
Embryonic Structures
Gene Knockout Techniques
Primates
Messenger RNA
Gene Editing
Oligonucleotides
Biomedical Research
Technology

ASJC Scopus subject areas

  • General

Cite this

Kumita, W., Sato, K., Suzuki, Y., Kurotaki, Y., Harada, T., Zhou, Y., ... Sasaki, E. (2019). Efficient generation of Knock-in/Knock-out marmoset embryo via CRISPR/Cas9 gene editing. Scientific reports, 9(1), [12719]. https://doi.org/10.1038/s41598-019-49110-3

Efficient generation of Knock-in/Knock-out marmoset embryo via CRISPR/Cas9 gene editing. / Kumita, Wakako; Sato, Kenya; Suzuki, Yasuhiro; Kurotaki, Yoko; Harada, Takeshi; Zhou, Yang; Kishi, Noriyuki; Sato, Kengo; Aiba, Atsu; Sakakibara, Yasubumi; Feng, Guoping; Okano, Hideyuki; Sasaki, Erika.

In: Scientific reports, Vol. 9, No. 1, 12719, 01.12.2019.

Research output: Contribution to journalArticle

Kumita, W, Sato, K, Suzuki, Y, Kurotaki, Y, Harada, T, Zhou, Y, Kishi, N, Sato, K, Aiba, A, Sakakibara, Y, Feng, G, Okano, H & Sasaki, E 2019, 'Efficient generation of Knock-in/Knock-out marmoset embryo via CRISPR/Cas9 gene editing', Scientific reports, vol. 9, no. 1, 12719. https://doi.org/10.1038/s41598-019-49110-3
Kumita, Wakako ; Sato, Kenya ; Suzuki, Yasuhiro ; Kurotaki, Yoko ; Harada, Takeshi ; Zhou, Yang ; Kishi, Noriyuki ; Sato, Kengo ; Aiba, Atsu ; Sakakibara, Yasubumi ; Feng, Guoping ; Okano, Hideyuki ; Sasaki, Erika. / Efficient generation of Knock-in/Knock-out marmoset embryo via CRISPR/Cas9 gene editing. In: Scientific reports. 2019 ; Vol. 9, No. 1.
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