title: High-throughput multiplex single-nucleotide polymorphism (SNP) analysis in genes involved in methionine metabolism.
abstract: Hyperhomocysteinemia is a well-known independent marker factor for atherothrombotic diseases and may result from acquired and genetic influences. Several polymorphisms are suspected to be associated with hyperhomocysteinemia, but data are limited and inconsistent. High-throughput genotyping technologies, such as GenomeLab SNPStream, are now available. Moreover, an appropriate selection of SNPs to be analyzed could represent a strong resource to define the role of genetic risk factors. We developed a multiplex PCR-oligonucleotide extension approach by GenomeLab platform. We selected 72 SNPs based on their putative function and frequency in the candidate genes AHCY, BHMT, BHMT2, CBS, ENOSF1, FOLH1, MTHFD1, MTHFR, MTR, MTRR, NNMT, PON1, PON2, SLC19A1, SHMT1, TCN2, and TYMS. We were able to analyze 57 of the SNPs (79%). For MTHFR C677T and A1298C and MTR A2756G SNPs, we compared data obtained with an electronic microchip technology and found 99.2% concordance. We also performed a haplotype analysis. This approach could represent a useful tool to investigate the genotype-phenotype correlation and the association of these genes with hyperhomocysteinemia and correlated diseases.
authors.affiliation: Department of Medical and Surgical Critical Care, University of Florence, Viale Morgagni 85, 50134, Florence, Italy. betti.giusti@unifi.it
authors.forename: Betti
authors.initials: B
authors.lastname: Giusti
authors.forename: Ilaria
authors.initials: I
authors.lastname: Sestini
authors.forename: Claudia
authors.initials: C
authors.lastname: Saracini
authors.forename: Elena
authors.initials: E
authors.lastname: Sticchi
authors.forename: Paola
authors.initials: P
authors.lastname: Bolli
authors.forename: Alberto
authors.initials: A
authors.lastname: Magi
authors.forename: Anna Maria
authors.initials: AM
authors.lastname: Gori
authors.forename: Rossella
authors.initials: R
authors.lastname: Marcucci
authors.forename: Gian Franco
authors.initials: GF
authors.lastname: Gensini
authors.forename: Rosanna
authors.initials: R
authors.lastname: Abbate
mesh_terms: D000328:Adult; D000368:Aged; D000369:Aged, 80 and over; D005260:Female; D005787:Gene Frequency; D006239:Haplotypes; D006801:Humans; D020138:Hyperhomocysteinemia / Q000235:genetics*; D015810:Linkage Disequilibrium; D008297:Male; D008715:Methionine / Q000378:metabolism*; D008875:Middle Aged; D020411:Oligonucleotide Array Sequence Analysis / Q000191:economics / Q000379:methods*; D016133:Polymerase Chain Reaction; D020641:Polymorphism, Single Nucleotide
chemical_list: D008715:Methionine