Bi-Allelic SNP Genotyping Using the TaqMan® Assay

Protocol
Part of the Methods in Molecular Biology book series (MIMB, volume 1145)

Abstract

With TaqMan® technology allele-specific probes are utilized for quick and reliable genotyping of known polymorphic sites. TaqMan assays are robust in genotyping multiple variant types, including single nucleotide polymorphisms, insertions/deletions, and presence/absence variants. To query a single bi-allelic polymorphism, two TaqMan probes labeled with distinct fluorophores are designed such that they hybridize to different alleles during PCR-based amplification of a surrounding target region. During the primer extension phase of PCR, the 5′–3′ exonuclease activity of Taq polymerase cleaves and releases the fluorophores from bound probes. At the end of PCR, the emission intensity of each fluorophore is measured and allele determination at the queried site can be made.

Key words

5′ Nuclease assay Applied biosystems 6-FAMTM VIC® Single nucleotide polymorphism Genotyping 

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Copyright information

© Springer Science+Business Media New York 2014

Authors and Affiliations

  1. 1.DuPont PioneerJohnstonUSA

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