A. Wavemaker-generated melting profiles for RET exon 10 without (top) and with (bottom) inclusion of a 36 base GC clamp on the 3′ amplification primer. The bold lines under the base pair axis indicate the extent of the primer sequences.
B. Predicted melting behavior of GC clamped RET10 amplicons at 65°C, 66°C, and 67°C. The GC clamp remains double stranded at all 3 temperatures. The target sequence is ≥ 90% double stranded at 65°C and 66°C, but not at 67°C.
C. CDHPLC elution profiles for 3 RET genotypes at 66°. Heterozygotes harboring the C620F and C620R mutations are readily distinguished from normals and from each other.