Smith J, Modrich P
Department of Biochemistry, Duke University Medical Center, Durham, NC 27710, USA.
Proc Natl Acad Sci U S A. 1996 Apr 30;93(9):4374-9. doi: 10.1073/pnas.93.9.4374.
Escherichia coli methyl-directed mismatch repair is initiated by MutS-, MutL-, and ATP-dependent activation of MutH endonuclease, which cleaves at d(GATC) sites in the vicinity of a mismatch. This reaction provides an efficient method for detection of mismatches in heteroduplexes produced by hybridization of genetically distinct sequences after PCR amplification. Multiple examples of transition and transversion mutations, as well as one, two, and three nucleotide insertion/deletion mutants, have been detected in PCR heteroduplexes ranging in size from 400 bp to 2.5 kb. Background cleavage of homoduplexes is largely due to polymerase errors that occur during amplification, and the MutHLS reaction provides an estimate of the incidence of mutant sequences that arise during PCR.
大肠杆菌甲基化导向错配修复由MutS、MutL和ATP依赖的MutH核酸内切酶激活引发，MutH核酸内切酶在错配附近的d(GATC)位点进行切割。该反应为检测PCR扩增后由遗传上不同的序列杂交产生的异源双链体中的错配提供了一种有效的方法。在大小从400 bp到2.5 kb的PCR异源双链体中，已检测到多个转换和颠换突变的例子，以及一、二和三个核苷酸插入/缺失突变体。同源双链体的背景切割主要是由于扩增过程中发生的聚合酶错误，而MutHLS反应提供了PCR过程中出现的突变序列发生率的估计值。
