Highly selective detection of single nucleotide polymorphism (SNP) using a dumbbell DNA probe with a gap-filling approach

Single nucleotide polymorphism (SNP) is the most common type of genetic sequence variations and known to associate with the development of various diseases including cancer. Accurate detection of SNP is often necessary to determine the susceptibility of patients to diseases as well as their response to the medical treatments. Molecular inversion probe (MIP) has been widely utilized as an alternative method to detect the SNP as compared to the conventional PCR/genome sequencing method. However, base-pairing based detection of SNP fundamentally lacks target specificity toward the single mismatch of bases and often fails to meet the accuracy requirement for SNP detection. In this study, we report the development of new SNP detection method using a dumbbell-probe with a gap-filling approach. Two separate DNA strands are designed to self-assemble to form a dumbbell DNA probe for the selective detection of EGFR 21 point mutation. Unlike the direct binding of MIP to the target sequence, our system has an additional process for the enhancement of sequence specificity through a base matched gap-filling. In the presence of target sequences, the dumbbell-probe undergoes base-pair hybridization to the target sequence followed by sequence specific gap-filling and ligation in single-step. The detection of EGFR 21 point mutation is achieved with a highly specific and selective manner. This simple and robust detection method offers promising future potential in the diagnosis of various genetic mutations.