FACS-assisted CRISPR-Cas9 genome editing of human induced pluripotent stem cells

被引：10

作者：

Caillaud, Amandine ^{[1
]}

Leveque, Antoine ^{[1
]}

Thedrez, Aurelie ^{[1
]}

Girardeau, Aurore ^{[1
]}

Canac, Robin ^{[1
]}

Bray, Lise ^{[1
]}

Baudic, Manon ^{[1
]}

Barc, Julien ^{[1
]}

Gaborit, Nathalie ^{[1
]}

Lamirault, Guillaume ^{[1
]}

Gardie, Betty ^{[1
]}

Idriss, Salam ^{[1
]}

Rimbert, Antoine ^{[1
]}

May, Cedric Le ^{[1
]}

Cariou, Bertrand ^{[1
,2
]}

Si-Tayeb, Karim ^{[1
]}

机构：

[1] Nantes Univ, CHU Nantes, INSERM, CNRS,Inst Thorax, F-44000 Nantes, France

[2] Univ Paris Sci & Lettres, Ecole Prat Hautes Etud, EPHE, Paris, France

来源：

STAR PROTOCOLS | 2022年 / 3卷 / 04期

关键词：

DELIVERY; PRECISE; FEEDER;

D O I：

10.1016/j.xpro.2022.101680

中图分类号：

Q5 [生物化学];

学科分类号：

071010 ; 081704 ;

摘要：

This manuscript proposes an efficient and reproducible protocol for the gener-ation of genetically modified human induced pluripotent stem cells (hiPSCs) by genome editing using CRISPR-Cas9 technology. Here, we describe the experi-mental strategy for generating knockout (KO) and knockin (KI) clonal popula-tions of hiPSCs using single-cell sorting by flow cytometry. We efficiently achieved up to 15 kb deletions, molecular tag insertions, and single-nucleotide editing in hiPSCs. We emphasize the efficacy of this approach in terms of cell culture time. For complete details on the use and execution of this protocol, please refer to Canac et al. (2022) and Bray et al. (2022).

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页数：19

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