Abstract
Many high-throughput single-nucleotide polymorphism (SNP) genotyping technologies are currently available. The method for SNP genotyping described in this unit is based on the commercially available Sequenom MassARRAY platform which offers several attractive features for users desiring an accurate SNP genotyping assay. The assay described is based on primer extension and offers two levels of specificity. First a locus-specific PCR reaction takes place, followed by a locus-specific primer extension reaction (homogeneous Mass Extend, or hME assay) in which an oligonucleotide primer anneals immediately upstream of the polymorphic site being genotyped. The extension of the primer is according to the sequence of the variant site and can be a single complementary base or a series of complementary bases. Through the use of MALDI-TOF mass spectrometry, the mass of the extended primer is determined. The preparation of probe and genomic DNA is also described in this unit.
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