ABSTRACT
Ever since the introduction of the LightCycler® in 1996 (Wittwer et al., 1997b), melting analysis has been an integral part of real-time techniques. SYBR® Green I allows both quantification without probes (Wittwer et al., 1997a) and verification of product identity by melting analysis (Ririe et al., 1997). However, fine sequence discrimination (e.g. SNP genotyping) has usually required labeled probes. Melting analysis for SNP typing was first achieved with one labeled primer and one labeled probe (Lay and Wittwer, 1997) and later with two labeled probes, each with a single label (Bernard et al., 1998). These techniques are widely used today with conventional realtime instrumentation (Wittwer and Kusukawa, 2004, Gingeras et al., 2005).
