High-throughput genotyping with infrared fluorescence allele specific hybridization (iFLASH): A simple, reliable and low-cost alternative

Abstract

Objectives To develop and validate a novel genotyping approach, named infrared Fluorescence Allele Specific Hybridization (iFLASH), which combines the principles of allele specific oligonucleotide (ASO) hybridization with the advanced possibilities of infrared imaging. Design and methods As an example, we genotyped the 55L > M and the 192Q > R common genetic variants of the paraoxonase-1 gene in 92 DNA samples using the iFLASH technique, and validated the outcomes with the restriction fragment length polymorphism (RFLP) and TAQman genotyping assays. Results There was a 100 percent agreement in genotype outcome among the three methods. Conclusions Although we found complete unity in genotype outcome, the iFLASH assay has essential advantages over the RFLP and TAQman genotyping assays. First, the iFLASH technique is capable of handling up to 1536 samples per assay, which makes it a suitable technique for high-throughput genotyping. Secondly, because the costs per assay are lower, high-throughput genotyping with iFLASH is affordable.