摘要
温度调控高效液相色谱法是探索人类基因组 DNA序列变异的新手段。温度调控高效液相色谱法通过比较异源双链与同源双链 ,而非逐个样本直接测序 ,可以敏感而准确地发现 DNA变异 ,唯发现的变异需进一步测序鉴定。对于旨在发现新突变或新多态性的课题 ,该方法可以明显减少测序工作量 ,而对于检出稀少的变异型 ,可明显降低所需成本。通过自动化操作 ,更能显著加快获取数据的速度。本文介绍温度调控高效液相色谱法的原理与命名。
Recently, temperature modulated heteroduplex analysis (TMHA) based upon ion pair reversed phase high performance liquid chromatography has been developed as a tool for polymorphisms and novel mutation detection in human genome.This new approach,with increased accuracy and throughput over traditional methods, is also known as denaturing high performance liquid chromatography (DHPLC). However, the terms of both TMHA and DHPLC should be considered carefully. Firstly, it is DNA molecular that is denatured in high performance liquid chromatography but not the method or the instrument. Secondly, the temperature in the liquid chromatograph is modulated according to Tm (melting temperature) of the DNA fragments. As it is known, Tm is the temperature at which a population of double stranded nucleic acid molecules becomes half dissociated into single strands,and under this temperature, half of the DNA molecules is still not denaturing. Thirdly, the targets for detection are both the homoduplexes and the mismatched heteroduplexes, which are double stranded nucleic acid molecules and are annealing. The term of DHPLC may lead to confusion in annealing and denaturing. In addition, since TMHA can be carried out using gel electrophoresis and HPLC, there is a need to distinguish HPLC from gel electrophoresis. Therefore, we suggest a new term for this technology, which is TmHPLC (temperature modulated high performance liquid chromatography). This term not only provides a clear definition for this new method, but also includes both TMHA and HPLC. More importantly, TmHPLC implies that the temperature in HPLC is modulated according to Tm of the DNA fragments. We also review advances in detecting variation through TmHPLC. This new approach leads to significant reduction of sequencing efforts for searching novel mutations and polymorphisms, and because of its high sample throughput, to faster data generation. This implies that TmHPLC is a sensitive, accurate, and cost effective approach to screening sequence variation in human
出处
《中华医学遗传学杂志》
EI
CAS
CSCD
北大核心
2000年第3期145-148,共4页
Chinese Journal of Medical Genetics
基金
国家自然科学金!(39993420)
关键词
温度调控
高效液相色谱
DAN变异
人类基因组
temperature modulated high performance liquid chromatography
denaturing high performance liquid chromatography
temperature modulated heteroduplex analysis
human genome
single nucleotide polymorphism
