Open AccessA duplex polymerase chain reaction‐restriction fragment length polymorphism for rapid screening of methylenetetrahydrofolate reductase gene variants: Genotyping in acute leukemia
Author(s) -
Frikha Rim,
Bouayed Nouha,
Ben Rhouma Bochra,
Keskes Leila,
Rebai Tarek
Publication year - 2018
Publication title -
journal of clinical laboratory analysis
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.536
H-Index - 50
eISSN - 1098-2825
pISSN - 0887-8013
DOI - 10.1002/jcla.22198
Subject(s) - methylenetetrahydrofolate reductase , genotyping , microbiology and biotechnology , polymerase chain reaction , restriction fragment length polymorphism , biology , amplified fragment length polymorphism , genomic dna , restriction enzyme , acute leukemia , dna extraction , genotype , genetics , leukemia , dna , gene , medicine , population , environmental health , genetic diversity
Background Methylenetetrahydrofolate reductase ( MTHFR ; NM _005957.4) is the key enzyme for folate metabolism which plays in DNA biosynthesis and the epigenetic process of DNA methylation. MTHFR gene polymorphisms, the c. 677C>T and c. 1298A>C have been implicated as risk factors for several types of cancers as the acute leukemia. Aim We have optimized a duplex polymerase chain reaction‐restriction fragment length polymorphism assay ( PCR ‐ RFLP ) for the simultaneous detection of both variants in acute leukemia patients, from Tunisia. Methods Genomic DNA was extracted from EDTA ‐anticoagulant blood samples from a total of 50 patients suffering from acute leukemia ( AL ). After DNA extraction, the polymerase chain reaction using specific primers, designed using Primer 3 Software. Restriction Fragment Length Polymorphism ( RFLP ) was performed in two separate tubes followed by agarose gel electrophoresis. Conclusion This new method has proved to be a rapid, simple, and reliable method that should facilitate high throughput genotyping of MTHFR polymorphisms in acute leukemia.