High-Throughput Single Nucleotide Polymorphism Genotyping by Fluorescent 5′ Exonuclease Assay

Single nucleotide polymorphism s (SNPs) are the class of DNA variants tha t are most common in the genome. Technolo gies are needed that allow relatively high throughput, high-quality genotyping o f SNPs, with rapid assay development for in dividual SNPs of interest. We have impl e mented an accurate and high-throughpu t SNP genotyping system using a comme r cially available fluorescent 5 ′ exonucleas e assay. Optimization of the assay system fo r low-volume reactions and low-probe con centrations has reduced assay costs b y 75%. Using a simple assay optimization process, we successfully developed genotyp ing assays for 92% of assays attempted (309 out of 335) and have generated ove r 200 000 genotypes .