Fig. 3

TaqMan duplex real-time PCR assay. a, b Amplification plots of wild-type (blue curve) and mutant (red curve) APC gene copies on synthetic wild-type and mutant DNAs (upper panels) and blood-derived DNAs of carrier or non-carrier JRTs (middle and lower panels). Amplification was plotted as fluorescence intensity (ΔRn value) against cycle numbers and shown on linear (a) and log (b) scales. The middle panels show representative results of individual cases of carrier or non-carrier dogs and the lower panels summarize the results of all examined cases (n = 11 and 5 for carrier and non-carrier JRTs, respectively). c Allelic discrimination plot based on the signal intensity ratio of FAM and VIC at the end points of PCR amplification. Synthetic wild-type and mutant DNAs are located at bottom right and top left corners, respectively, and distilled water used as negative control is at bottom left corner. Non-carrier JRTs are clustered together with the synthetic wild-type DNA at bottom right corner. Carrier JRTs are located near the diagonal line forming a distinct cluster