Specific and efficient gene knockout and overexpression in mouse interscapular brown adipocytes in vivo

Kaili Xue; Dongmei Wu; Yifu Qiu

doi:10.1016/j.xpro.2022.101895

Specific and efficient gene knockout and overexpression in mouse interscapular brown adipocytes in vivo

STAR Protoc. 2022 Dec 16;3(4):101895. doi: 10.1016/j.xpro.2022.101895. Epub 2022 Dec 2.

Authors

Kaili Xue¹, Dongmei Wu², Yifu Qiu³

Affiliations

¹ Institute of Molecular Medicine, Beijing Key Laboratory of Cardiometabolic Molecular Medicine, College of Future Technology, Peking University, Beijing 100871, China. Electronic address: kaili.xue@pku.edu.cn.
² Institute of Molecular Medicine, Beijing Key Laboratory of Cardiometabolic Molecular Medicine, College of Future Technology, Peking University, Beijing 100871, China; Peking-Tsinghua Center for Life Sciences, Peking University, Beijing 100871, China.
³ Institute of Molecular Medicine, Beijing Key Laboratory of Cardiometabolic Molecular Medicine, College of Future Technology, Peking University, Beijing 100871, China; Peking-Tsinghua Center for Life Sciences, Peking University, Beijing 100871, China. Electronic address: yifu.qiu@pku.edu.cn.

Abstract

The classical Cre-LoxP system is time consuming. Here we detail a protocol that leverages Rosa26-LSL-Cas9;Adiponectin-Cre mice to restrict Cas9 expression in adipocytes. This enables specific deletion of target genes in brown adipocytes within 6 weeks by local injection of AAV-sgRNA into interscapular brown adipose tissue. We also describe an adiponectin-promoter-driven AAV vector to express sgRNA-resistant cDNA-encoded protein for subsequent rescue. This protocol thus provides an efficient means to specifically knockout and overexpress genes in brown adipocytes in vivo. For complete details on the use and execution of this protocol, please refer to Xue et al. (2022).¹.

Keywords: CRISPR; Metabolism; Model Organisms; Molecular Biology.

Publication types

Research Support, Non-U.S. Gov't

MeSH terms

Adipocytes, Brown* / metabolism
Adiponectin* / genetics
Adiponectin* / metabolism
Adipose Tissue, Brown / metabolism
Animals
Gene Knockout Techniques
Mice
Promoter Regions, Genetic

Substances

Adiponectin