Temperature-modulated high-performance liquid chromatography for detecting variation in human genome.
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. This new approach 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 genome.
Hou
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Y
Zhang
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S
D002851:Chromatography, High Pressure Liquid; D014644:Genetic Variation; D015894:Genome, Human; D020180:Heteroduplex Analysis; D006801:Humans; D011110:Polymorphism, Genetic; D013696:Temperature