Genotyping of Single-Nucleotide Polymorphisms by High-Resolution Melting of Small Amplicons

cons with the DNA dye LCGreenTM I was recently introduced as a homogeneous, closed-tube method of genotyping that does not require probes or real-time PCR. We adapted this system to genotype single-nucle-otide polymorphisms (SNPs) after rapid-cycle PCR (12 min) of small amplicons (<50 bp). Methods: Engineered plasmids were used to study all possible SNP base changes. In addition, clinical proto-cols for factor V (Leiden) 1691G>A, prothrombin 20210G>A, methylenetetrahydrofolate reductase (MTHFR) 1298A>C, hemochromatosis (HFE) 187C>G, and -glo-bin (hemoglobin S) 17A>T were developed. LCGreen I was included in the reaction mixture before PCR, and high-resolution melting was obtained within 2 min after amplification