Chen Huan, Liu Xiang-Jun, Liu Ya-Li, Jiang Jian-Hui, Shen Guo-Li, Yu Ru-Qin
State Key Laboratory for Chemo/Biosensing and Chemometrics, College of Chemistry and Chemical Engineering, Hunan University, Yuelu District, Changsha 410082, PR China.
Biosens Bioelectron. 2009 Mar 15;24(7):1955-61. doi: 10.1016/j.bios.2008.09.029. Epub 2008 Oct 15.
MutS protein is an important part of the DNA repair system which can specifically recognize and bind all possible single-base mismatches as well as 1-4 base insertion or deletion loops with varying affinities independent of other proteins or cofactors. In this paper, a new approach for electrochemical gene mutation detection based on the utilization of MutS protein for the mutation recognition and spontaneously intercalated methylene blue (MB) markers for electrochemical signal generation is described. This method involves the immobilization of MutS protein onto the gold electrode, the hybridization of target DNA to form homoduplex or heteroduplex DNA, the application of MutS protein for the mutation recognition, and finally the intercalation of MB. The background is very low because MutS protein binds DNA containing mispaired and unpaired bases but does not bind equally well to DNA without mismatches or single-stranded DNA. The proposed approach has been successfully implemented for the identification of single-base mutation in -28 site of the beta-thalassemia gene with a detection limit of 5.6 x 10(-13)M, demonstrating that this method provides a highly specific and cost-efficient approach for point mutation detection.
MutS蛋白是DNA修复系统的重要组成部分，它能够特异性识别并结合所有可能的单碱基错配以及1至4个碱基的插入或缺失环，且结合亲和力各异，无需其他蛋白质或辅助因子。本文描述了一种基于利用MutS蛋白进行突变识别以及利用自发插入的亚甲基蓝（MB）标记物产生电化学信号的电化学基因突变检测新方法。该方法包括将MutS蛋白固定在金电极上，使目标DNA杂交形成同源双链或异源双链DNA，应用MutS蛋白进行突变识别，最后插入MB。背景信号非常低，因为MutS蛋白能结合含有错配和未配对碱基的DNA，但对无错配的DNA或单链DNA的结合能力不同。所提出的方法已成功用于鉴定β地中海贫血基因-28位点的单碱基突变，检测限为5.6×10（-13）M，表明该方法为点突变检测提供了一种高度特异且经济高效的方法。
