Microarray-based method for genotyping of functional single nucleotide polymorphisms using dual-color fluorescence hybridization.
Screening disease-related single nucleotide polymorphism (SNP) markers in the whole genome has great potential in complex disease genetics and pharmacogenetics researches. It has led to a requirement for high-throughput genotyping platforms that can maximize the efficient screening functional SNPs with respect to accuracy, speed and cost. In this study, we attempted to develop a microarray-based method for scoring a number of genomic DNA in parallel for one or more molecular markers on a glass slide. Two SNP markers localized to the methylenetetrahydrofolate reductase gene (MTHFR) were selected as the investigated targets. Amplified PCR products from nine genomic DNA specimens were spotted and immobilized onto a poly-l-lysine coated glass slide to fabricate a microarray, then interrogated by hybridization with dual-color probes to determine the SNP genotype of each sample. The results indicated that the microarray-based method could determine the genotype of 677 and 1298 MTHFR polymorphisms. Sequencing was performed to validate these results. Our experiments successfully demonstrate that PCR products subjected to dual-color hybridization on a microarray could be applied as a useful and a high-throughput tool to analyze molecular markers.
Ji
Meiju
M
Chien-Shiung Wu Laboratory, Department of Biological Science and Medical Engineering, Southeast University, Nanjing 210096, China.
Hou
Peng
P
Li
Song
S
He
Nongyue
N
Lu
Zuhong
Z
D005456:Fluorescent Dyes; D005838:Genotype; D006801:Humans; D006824:Hybridization, Genetic; D042965:Methylenetetrahydrofolate Reductase (NADPH2) / Q000235:genetics*; D009303:Nasopharyngeal Neoplasms / Q000235:genetics*; D020411:Oligonucleotide Array Sequence Analysis / Q000379:methods*; D016133:Polymerase Chain Reaction / Q000379:methods; D020641:Polymorphism, Single Nucleotide; D017422:Sequence Analysis, DNA
D005456:Fluorescent Dyes; D042965:Methylenetetrahydrofolate Reductase (NADPH2)