Detection of C677T mutation in methylenetetrahydrofolate reductase gene by denaturing high performance liquid chromatography.
In this paper, we described an assay for the detection of the C677T mutation in the methylenetetrahydrofolate reductase (MTHFR) gene using denaturing high-performance liquid chromatography (DHPLC). The conditions for DHPLC analysis were systematically investigated based on a general HPLC instrument (Prostar VARIAN). A 225 bp DNA fragment covering the 677 site of MTHFR gene was amplified by PCR technology using the purified DNA from whole blood or whole blood as template DNA. PCR products were directly injected without the need for purification. The C677T mutation could be clearly distinguished by DHPLC technology. Our data demonstrated that DHPLC was a powerful and alternative tool for detection of genetic variants and single-nucleotide polymorphisms to electrophoresis technology.
Fang
Nenghu
N
Department of Chemistry, Shanghai Jiaotong University, Shanghai, 200240, People's Republic of China. nhfang@sjtu.edu.cn
Lin
Li
L
Ren
Jicun
J
Wu
Dan
D
D001483:Base Sequence; D002851:Chromatography, High Pressure Liquid / Q000379:methods*; D017931:DNA Primers; D042965:Methylenetetrahydrofolate Reductase (NADPH2) / Q000737:chemistry / Q000235:genetics*; D009154:Mutation; D016133:Polymerase Chain Reaction; D011489:Protein Denaturation
D017931:DNA Primers; D042965:Methylenetetrahydrofolate Reductase (NADPH2)