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Genotyping of single nucleotide polymorphism in γ‐glutamyl hydrolase gene by capillary electrophoresis
Author(s) -
Cheng HuiLing,
Chiou ShyhShin,
Liao YuMei,
Chen YenLing,
Wu ShouMei
Publication year - 2011
Publication title -
electrophoresis
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.666
H-Index - 158
eISSN - 1522-2683
pISSN - 0173-0835
DOI - 10.1002/elps.201000422
Subject(s) - genotyping , single nucleotide polymorphism , microbiology and biotechnology , capillary electrophoresis , snp , genotype , chemistry , genomic dna , gene , single strand conformation polymorphism , dna , lymphoblastic leukemia , biology , exon , biochemistry , leukemia , genetics
The γ‐glutamyl hydrolase ( GGH ) gene plays an important role in methotrexate (MTX) metabolism, ensuring that MTX polyglutamates (MTX‐(Glu) n ) could be converted back into MTX. Accumulation of MTX‐(Glu) n is a problem in MTX therapy. SNP 452 C>T has been reported to associate with lower catalytic activity and higher accumulation of long‐chain MTX‐(Glu) n in patients treated with higher doses of MTX treatment. We propose and establish a simple and effective CE method for detecting SNP in GGH gene. The DNA samples after amplification were analyzed by SSCP‐CE method. The CE conditions were generated by using 1× TBE buffer containing 1.5% w/v hydroxypropyl methyl cellulose under reverse polarity at 25°C. This method was applied to detect genotyping of acute lymphoblastic leukemia patients receiving MTX treatment. The results were confirmed by DNA sequencing with good agreement. Concentrations of MTX‐(Glu) n in whole blood were analyzed by on‐line stacking CE method. MTX‐(Glu) n levels and genotypes in GGH gene of acute lymphoblastic leukemia patients were evaluated. The SSCP‐CE method was found to be feasible for SNP screening in the GGH gene.