2-TEMPERATURE PCR AND HETERODUPLEX DETECTION - APPLICATION TO RAPID CYSTIC-FIBROSIS SCREENING

We describe a rapid two-temperature PCR protocol for amplification of genomic DNA applied to the region of the most common mutation (ΔF508) of the cystic fibrosis gene. Amplification products are detected as homo- or heteroduplexes on polyacrylamide gels as previously described.1 Data using two-temperature PCR show complete concordance with allele-specific hybridization after classical three-temperature PCR in 105 normal, carrier and affected individuals. Clinical application is demonstrated in a family which was uninformative by traditional RFLP linkage analysis. Two-temperature PCR may offer advantages of speed and specificity over three-temperature PCR in many clinical and research applications. © 1991.