Figure 2.

cDNA analysis and haplotype reconstruction. (A) CFTR cDNA spanning the regions containing the variants was amplified by RT-PCR and the obtained products checked by Sanger sequencing, thus disclosing the presence of a third substitution, the L467F. As shown, we detected an allelic imbalance due to NMD mRNA degradation of the G542X-carrying transcript with the corresponding sequence peaks less intense compared to those showing the presence of the L467F and F508del variants, thus indicating that these latter are in cis and configure a complex allele. (B) Schematic representation of the allelic phase definition. The complex allele is also carrying the common V470M polymorphism frequently observed associated with pathogenic CFTR variants. A1, allele 1; A2, allele 2; NMD, nonsense-mediated mRNA decay.