SNP Genotyping by the 5-Nuclease Reaction: Advances in High-Throughput Genotyping with Nonmodel Organisms

Population genetics studies play an increasingly important role in the management and conservation of nonmodel organisms. Unlike studies with model organisms, a typical population genetics study of a nonmodel organism may be conducted by analyzing thousands or hundreds of thousands of individuals for several dozen single nucleotide polymorphisms (SNPs). The use of robust, robotically mediated TaqMan reactions provides substantial advantages in these types of studies. We describe the methods and laboratory setup for analyzing a sustained high throughput of SNP assays in routine university or natural resource agency laboratories with a handful of thermal cyclers. Agencies sustain rates of nearly 150,000 assays per week using uniplex reactions with the Applied Biosystems 7900HT Fast Real-Time PCR System (AB 7900HT). We further describe the medium-density array run on the BioMark from Fluidigm, which increases this rate to over 500,000 assays per week by multiplexing 96 samples for 96 SNPs.