Protocol for genetic dissection of class switch recombination using genome-editing tools

Pengfei Dai; Tingting Liu; Dingpeng Yang; Yifeng Luo; Liu Daisy Liu; Xia Xie; Yafang Shang; Xiaojing Liu; Fei-Long Meng

STAR Protocols (Jun 2025)

Protocol for genetic dissection of class switch recombination using genome-editing tools

Pengfei Dai,
Tingting Liu,
Dingpeng Yang,
Yifeng Luo,
Liu Daisy Liu,
Xia Xie,
Yafang Shang,
Xiaojing Liu,
Fei-Long Meng

Affiliations

Pengfei Dai: Key Laboratory of RNA Innovation, Science and Engineering, Shanghai Academy of Natural Sciences (SANS), Shanghai Institute of Biochemistry and Cell Biology, Center for Excellence in Molecular Cell Science, Chinese Academy of Sciences, University of Chinese Academy of Sciences, Shanghai 200031, China; International Peace Maternity and Child Health Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai Key Laboratory of Embryo Original Diseases, Shanghai 200030, China; Corresponding author
Tingting Liu: Key Laboratory of RNA Innovation, Science and Engineering, Shanghai Academy of Natural Sciences (SANS), Shanghai Institute of Biochemistry and Cell Biology, Center for Excellence in Molecular Cell Science, Chinese Academy of Sciences, University of Chinese Academy of Sciences, Shanghai 200031, China
Dingpeng Yang: Key Laboratory of RNA Innovation, Science and Engineering, Shanghai Academy of Natural Sciences (SANS), Shanghai Institute of Biochemistry and Cell Biology, Center for Excellence in Molecular Cell Science, Chinese Academy of Sciences, University of Chinese Academy of Sciences, Shanghai 200031, China
Yifeng Luo: Key Laboratory of RNA Innovation, Science and Engineering, Shanghai Academy of Natural Sciences (SANS), Shanghai Institute of Biochemistry and Cell Biology, Center for Excellence in Molecular Cell Science, Chinese Academy of Sciences, University of Chinese Academy of Sciences, Shanghai 200031, China
Liu Daisy Liu: Key Laboratory of RNA Innovation, Science and Engineering, Shanghai Academy of Natural Sciences (SANS), Shanghai Institute of Biochemistry and Cell Biology, Center for Excellence in Molecular Cell Science, Chinese Academy of Sciences, University of Chinese Academy of Sciences, Shanghai 200031, China
Xia Xie: Key Laboratory of RNA Innovation, Science and Engineering, Shanghai Academy of Natural Sciences (SANS), Shanghai Institute of Biochemistry and Cell Biology, Center for Excellence in Molecular Cell Science, Chinese Academy of Sciences, University of Chinese Academy of Sciences, Shanghai 200031, China
Yafang Shang: Key Laboratory of RNA Innovation, Science and Engineering, Shanghai Academy of Natural Sciences (SANS), Shanghai Institute of Biochemistry and Cell Biology, Center for Excellence in Molecular Cell Science, Chinese Academy of Sciences, University of Chinese Academy of Sciences, Shanghai 200031, China
Xiaojing Liu: Key Laboratory of RNA Innovation, Science and Engineering, Shanghai Academy of Natural Sciences (SANS), Shanghai Institute of Biochemistry and Cell Biology, Center for Excellence in Molecular Cell Science, Chinese Academy of Sciences, University of Chinese Academy of Sciences, Shanghai 200031, China
Fei-Long Meng: Key Laboratory of RNA Innovation, Science and Engineering, Shanghai Academy of Natural Sciences (SANS), Shanghai Institute of Biochemistry and Cell Biology, Center for Excellence in Molecular Cell Science, Chinese Academy of Sciences, University of Chinese Academy of Sciences, Shanghai 200031, China; Corresponding author

Journal volume & issue: Vol. 6, no. 2
p. 103882

Abstract

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Summary: Antibody class switch recombination is achieved through programmed DNA damage, and the processing of programmed DNA lesions requires the coordinated action of many DNA metabolic factors. Here, we present a protocol for inducing class switch recombination using base editors or CRISPR-Cas9. We provide optimized guide RNA (gRNA) sequences and describe steps for cytokine activation, electroporation, surface immunoglobulin detection, and data analysis. This method allows researchers to investigate the involvement of specific factors in antibody diversification and elucidate their functional roles.For complete details on the use and execution of this protocol, please refer to Dai et al.1 : Publisher’s note: Undertaking any experimental protocol requires adherence to local institutional guidelines for laboratory safety and ethics.

Published in STAR Protocols

ISSN: 2666-1667 (Online)
Publisher: Elsevier
Country of publisher: United States
LCC subjects: Science: Science (General)
Website: https://star-protocols.cell.com/

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