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利用自动化电生理学技术对人类离子通道变体进行功能评估。

Functional evaluation of human ion channel variants using automated electrophysiology.

机构信息

Department of Pharmacology, Northwestern University Feinberg School of Medicine, Chicago, IL, United States.

Department of Preventive Medicine, Northwestern University Feinberg School of Medicine, Chicago, IL, United States.

出版信息

Methods Enzymol. 2021;654:383-405. doi: 10.1016/bs.mie.2021.02.011. Epub 2021 Apr 9.

Abstract

Patch clamp recording enabled a revolution in cellular electrophysiology, and is useful for evaluating the functional consequences of ion channel gene mutations or variants associated with human disorders called channelopathies. However, due to massive growth of genetic testing in medical practice and research, the number of known ion channel variants has exploded into the thousands. Fortunately, automated methods for performing patch clamp recording have emerged as important tools to address the explosion in ion channel variants. In this chapter, we present our approach to harnessing automated electrophysiology to study a human voltage-gated potassium channel gene (KCNQ1), which harbors hundreds of mutations associated with genetic disorders of heart rhythm including the congenital long-QT syndrome. We include protocols for performing high efficiency electroporation of heterologous cells with recombinant KCNQ1 plasmid DNA and for automated planar patch recording including data analysis. These methods can be adapted for studying other voltage-gated ion channels.

摘要

膜片钳记录技术引发了细胞电生理学的革命,对于评估与人类疾病(称为通道病)相关的离子通道基因突变或变体的功能后果非常有用。然而,由于医学实践和研究中基因检测数量的大量增加,已知的离子通道变体数量已经爆炸式增长到数千种。幸运的是,自动化膜片钳记录方法已成为研究离子通道变体的重要工具。在本章中,我们介绍了利用自动化电生理学研究人类电压门控钾通道基因(KCNQ1)的方法,该基因携带有数百种与心律失常遗传疾病相关的突变,包括先天性长 QT 综合征。我们包括了用重组 KCNQ1 质粒 DNA 高效转染异源细胞的方案以及自动化平面膜片钳记录(包括数据分析)的方案。这些方法可以适用于研究其他电压门控离子通道。

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