利用基于游离型载体的CRISPR/Cas9建立TRPV2基因敲除的人胚胎干细胞系（WAe009-A-1Y）

Establishment of a TRPV2 knockout human embryonic stem cell line (WAe009-A-1Y) using episomal vector-based CRISPR/Cas9.

作者信息

Bai Rui, Zhang Siyao, Gu Xiaoyan, You Yi, Liu Xujie

机构信息

State Key Laboratory of Cardiovascular Disease, Fuwai Hospital Chinese Academy of Medical Sciences, Shenzhen, Guangdong Province, China.

Maternal-Fetal Medicine Center in Fetal Heart Disease, Capital Medical University, Beijing Anzhen Hospital, Beijing 100029, China; Beijing Lab for Cardiovascular Precision Medicine, Capital Medical University, No. 2 Anzhen Road, Chaoyang District, Beijing 100069, China.

出版信息

Stem Cell Res. 2025 Sep;87:103744. doi: 10.1016/j.scr.2025.103744. Epub 2025 May 30.

DOI:10.1016/j.scr.2025.103744

PMID:40483902

Abstract

We established a TRPV2-knockout human embryonic stem cell line (WAe009-A-1Y) using a non-integrating episomal CRISPR/Cas9 system. This cell line exhibits an 8-nucleotide frameshift deletion in TRPV2 exon 2, confirmed pluripotency (97.6 % SSEA4+ cells, trilineage differentiation), and a normal female karyotype (46, XX) at passage 30. TRPV2 ablation was validated in differentiated cardiomyocytes, showing >90 % mRNA reduction and absent protein expression. No off-target edits or mycoplasma contamination were detected. This cell resource (STR-authenticated, off-target-free) provides a robust in vitro model to study the biological function of TRPV2 in cardiac mechanotransduction and disease.

摘要

我们使用非整合型附加体CRISPR/Cas9系统建立了一个TRPV2基因敲除的人胚胎干细胞系（WAe009-A-1Y）。该细胞系在TRPV2外显子2中表现出8个核苷酸的移码缺失，在第30代时证实具有多能性（97.6%的细胞SSEA4呈阳性，具有三系分化能力），且核型为正常女性核型（46, XX）。在分化的心肌细胞中验证了TRPV2基因的缺失，结果显示mRNA减少超过90%且无蛋白表达。未检测到脱靶编辑或支原体污染。这种细胞资源（经过STR鉴定且无脱靶）为研究TRPV2在心脏机械转导和疾病中的生物学功能提供了一个强大的体外模型。