title: An unusual melting curve on a LightCycler due to a new mutation in methylenetetrahydrofolate reductase.
abstract: Methylenetetrahydrofolate reductase (MTHFR; E.C. 1.5.1.20) catalyzes the reduction of 5,10-methylenetetrahydofolate to 5-methytetrahydrofolate, and its activity is affected by a number of polymorphisms and mutations, the best known being MTHFR c.677C > T (OMIM 607093.003). The c.677C > T mutation is often analyzed by melting curve analysis after real-time polymerase chain reaction. We observed in one patient upon dual hybridization probe analysis in a Roche LightCycler 1.2 an unusual melting curve peak at 53.4 degrees C instead of one expected at either 51.6 degrees C or 60.4 degrees C. We sequenced the region of MTHFR interrogated by the dual hybridization probes, which revealed a c.685A > G base substitution that would result in an Ile225Val substitution. In conclusion, we identified the cause of an unusual melting curve in a commonly used genotyping technique for MTHFR c.677C > T as due to a novel mutation highlighting the ability of melting analysis to identify new sequence variants.
authors.affiliation: Division of Medical Biochemistry, McGill University Health Centre (MUHC)-Royal Victoria Hospital, Montreal, Canada. brian.gilfix@mcgill.ca
authors.forename: Brian M
authors.initials: BM
authors.lastname: Gilfix
mesh_terms: D000328:Adult; D019943:Amino Acid Substitution; D004247:DNA / Q000737:chemistry* / Q000235:genetics; D015342:DNA Probes / Q000737:chemistry / Q000235:genetics; D005260:Female; D005820:Genetic Testing; D006801:Humans; D007532:Isoleucine / Q000235:genetics; D007930:Leucine / Q000235:genetics; D042965:Methylenetetrahydrofolate Reductase (NADPH2) / Q000235:genetics*; D009154:Mutation; D009693:Nucleic Acid Hybridization; D016133:Polymerase Chain Reaction; D044366:Transition Temperature
chemical_list: D015342:DNA Probes; D007532:Isoleucine; D004247:DNA; D042965:Methylenetetrahydrofolate Reductase (NADPH2); D007930:Leucine