1. A Simple and Rapid Genotyping Assay for Simultaneous Detection of Two ADRB2 Allelic Variants Using Fluorescence Resonance Energy Transfer Probes and Melting Curve Analysis 
M. Fernanda Sábato, Anne-Marie Irani, Bonny L. Bukaveckas, Lawrence B. Schwartz, David S. Wilkinson, Andrea Ferreira-Gonzalez 
The Journal of Molecular Diagnostics 
Volume 10, Issue 3, Pages (May 2008)
DOI: /jmoldx
Copyright © 2008 American Society for Investigative Pathology and Association for Molecular Pathology Terms and Conditions

2. Figure 1
Diagram showing positions of primers and hybridization probes for the ADRB2 genotyping assay. An amplicon that spans the area of the allelic variants is created with one set of primers. A single anchor probe and two sensor probes anneal to the forward strand internal to the primer set. During FRET, the long anchor probe labeled at the 5′ and 3′ ends with fluorescein is excited by the light source of the LightCycler instrument and then passes on part of its excitation energy via dipole-dipole interactions to the adjacent acceptors, the LC-Red 705 and LC-Red 640 in the so-called sensor probes that cover the allelic variants at codons 16 and 27 of the β-2-adrenergic receptor, respectively. The excited fluorophores emit measurable light in channels 3 and 2 of the LightCycler instrument, respectively. The DNA sequence changes (c.46A>G and c.79C>G) are shown in capital letters.
The Journal of Molecular Diagnostics  , DOI: ( /jmoldx )
Copyright © 2008 American Society for Investigative Pathology and Association for Molecular Pathology Terms and Conditions

3. Figure 2
Melting curves and melting peaks for each sensor probe. One DNA representing each genotype was used to optimize PCR conditions and LightCycler protocol as described in Materials and Methods. Amplicons were denatured and the probes melted at the rate of 0.1°C/second. A: Melting curves (top) and derivative melting peaks (bottom) for genotypes detected by the LC Red-705 sensor probe. The asterisk represents an unexplained elevation (shoulder) during the melting analysis of the p.Gly16Gly genotype. B: Melting curves (top) and derivative melting peaks (bottom) for genotypes detected by the LC Red-640 sensor probe.
The Journal of Molecular Diagnostics  , DOI: ( /jmoldx )
Copyright © 2008 American Society for Investigative Pathology and Association for Molecular Pathology Terms and Conditions