Generation of INS-jGCaMP7f knock-in Ca2+ reporter human embryonic stem cell line, GZLe001-C, using CRISPR/Cas9-based gene targeting

Xin Liu; Feng Zhang; Deqi Chen; Jiaxiang Yin; Zirong Bi; Lihua Chen; Jiawei Yan; Qifei Dong; Wei Peng; Tao Xu; Yanying Guo; Haopeng Lin; Huisheng Liu

doi:10.1016/j.scr.2024.103633

Generation of INS-jGCaMP7f knock-in Ca²⁺ reporter human embryonic stem cell line, GZLe001-C, using CRISPR/Cas9-based gene targeting

Stem Cell Res. 2024 Dec 18:82:103633. doi: 10.1016/j.scr.2024.103633. Online ahead of print.

Authors

Xin Liu¹, Feng Zhang², Deqi Chen², Jiaxiang Yin³, Zirong Bi², Lihua Chen³, Jiawei Yan³, Qifei Dong⁴, Wei Peng⁴, Tao Xu³, Yanying Guo⁵, Haopeng Lin³, Huisheng Liu⁶

Affiliations

¹ Department of Endocrinology and Metabolism, People's Hospital of Xinjiang Uygur Autonomous Region, Xinjiang Clinical Research Center for Diabetes, Xinjiang Key Laboratory of Cardiovascular Homeostasis and Regeneration Research, Urumqi 830001, China; Guangzhou National Laboratory, Guangzhou 510005, China.
² Guangzhou National Laboratory, Guangzhou 510005, China.
³ Guangzhou National Laboratory, Guangzhou 510005, China; School of Biomedical Engineering, Guangzhou Medical University, Guangzhou 510180, China.
⁴ Guangzhou National Laboratory, Guangzhou 510005, China; Institute of Cytology and Genetics, School of Basic Medical Sciences, Hengyang Medical School, University of South China, Hengyang, Hunan 421001, China.
⁵ Department of Endocrinology and Metabolism, People's Hospital of Xinjiang Uygur Autonomous Region, Xinjiang Clinical Research Center for Diabetes, Xinjiang Key Laboratory of Cardiovascular Homeostasis and Regeneration Research, Urumqi 830001, China.
⁶ Guangzhou National Laboratory, Guangzhou 510005, China; School of Biomedical Engineering, Guangzhou Medical University, Guangzhou 510180, China; Bioland Laboratory, Guangzhou 510005, China.

PMID: 39708407
DOI: 10.1016/j.scr.2024.103633

Abstract

As a member of the single-fluorophore genetically encoded calcium indicators (GECIs), jGCaMP7f is widely applied to investigate intracellular Ca²⁺ concentrations. Here, we established an INS-jGCaMP7f knock-in H1 human embryonic stem cell (hESC) line by integrating jGCaMP7f gene into insulin locus via CRISPR/Cas9 system. The reporter cell line not only effectively labelled the insulin-producing cells induced from hESC, but also reflected the cytosolic change of Ca²⁺ level in response to different stimuli. This reporter cell line is a valuable research tool for studying functional maturation of hESC-derived insulin-producing cells, conducting drug screenings, and exploring the mechanisms of diabetes.

Keywords: CRISPR/Cas9; Ca(2+); Insulin; hESC; jGCaMP7f.