从细胞和组织中定量全细胞和线粒体 dNTP 池。

Whole-Cell and Mitochondrial dNTP Pool Quantification from Cells and Tissues.

机构信息

Research Programs Unit, Stem Cells and Metabolism, University of Helsinki, Helsinki, Finland.

Department of Anatomy, Physiology and Biochemistry, Swedish University of Agricultural Sciences, Uppsala, Sweden.

出版信息

Methods Mol Biol. 2021;2276:143-151. doi: 10.1007/978-1-0716-1266-8_10.

DOI:10.1007/978-1-0716-1266-8_10

PMID:34060038

Abstract

Deoxynucleoside 5'-triphosphates (dNTPs) are the molecular building blocks for DNA synthesis, and their balanced concentration in the cell is fundamental for health. dNTP imbalance can lead to genomic instability and other metabolic disturbances, resulting in devastating mitochondrial diseases.The accurate and efficient measurement of dNTPs from different biological samples and cellular compartments is vital to understand the mechanisms behind these diseases and develop and scrutinize their possible treatments. This chapter describes an update on the most recent development of the traditional radiolabeled polymerase extension method and its adaptation for the measurement of whole-cell and mitochondrial dNTP pools from cultured cells and tissue samples. The solid-phase reaction setting enables an increase in efficiency, accuracy, and measurement scale.

摘要

脱氧核苷 5'-三磷酸 (dNTPs) 是 DNA 合成的分子构建块，其在细胞中的平衡浓度对健康至关重要。dNTP 失衡可导致基因组不稳定和其他代谢紊乱，从而导致毁灭性的线粒体疾病。准确、有效地测量来自不同生物样本和细胞区室的 dNTP 对于理解这些疾病背后的机制以及开发和审查其可能的治疗方法至关重要。本章介绍了传统放射性标记聚合酶延伸方法的最新进展及其适应从培养细胞和组织样本中测量全细胞和线粒体 dNTP 池的方法。固相反应设置提高了效率、准确性和测量规模。