Detection of single nucleotide polymorphisms (SNPs) is of great value in precision medicine. The polymorphism of the aldehyde dehydrogenase 2 (ALDH2) gene is caused by a G1510A transition, resulting in the substitution of glutamic acid by lysine at position 487. People of different ALDH2 genotypes show different susceptibility to cancer, metabolic diseases, etc. SNP analysis based on fluorescent probe-mediated melting curves is a relatively efficient and cost-effective method. Genomic DNA extracted from 100 whole blood samples was subjected to polymorphisms mutational analysis using asymmetric PCR and probe-mediated melting curves. Then a certain number of samples from each genotype were randomly selected for direct sequencing verification. The new assay can be performed in 2 h without post-PCR processing such as gel electrophoresis and validated by direct sequencing in a blind study with 100% concordance. Moreover, comparing the detection of polymorphisms of ALDH2 with the clinics, and an overall agreement of 100% (100/100) was demonstrated. Our study has shown a high level of concordance between DNA sequencing, which is suitable for the detection of clinical specimens. Based on the concept of probe-mediated melting curves, we further developed this platform as a universal strategy for the detection of polymorphisms related to folate metabolism. In this paper, based on asymmetric PCR and probe-mediated melting curves method is proposed to detect ALDH2 polymorphism. As shown in graphical abstract, the genomic DNA in whole blood sample was extracted by Genomic DNA Extraction Kit and underwent an asymmetric PCR to enrich the target sequence by designed primers. During the asymmetric PCR, the excess primer (yellow) generates excess copies of single-strand amplicons, then during the melting analysis, the probe binds to the single-stranded amplicons at lower temperatures to form a hybridized double strand, and as the temperature increases during the melting analysis process, the probe gradually dissociates from the target strand and the fluorescence signal decreases because of FRET. The differences of a single base pair will be recorded on the melting temperature due to the short probe length. [Display omitted] • A newly-developed detection platform was reported by combining asymmetric PCR and fluorescent probe-mediated melting curves. • The method exhibited excellent accuracy to detect polymorphisms in real samples. • The analysis showed high sensitivity with the LOD of 5 ng/μL for correct results interpretation. • The proposed strategy achieved universality in polymorphism detection for other loci. [ABSTRACT FROM AUTHOR]