Eighteen Hb Lepore carriers from six unrelated families were studied. Hematological parameters were obtained using an automated cell counter. Hb F was quantitated by standard methods [11,12] and Hb A₂ was estimated by elution from celogel electrophoretic strips. The abnormal Hb was detected by electrophoresis on cellulose acetate in Tris-borate-EDTA (pH 8.4) [13]. Genomic DNA was extracted from peripheral blood collected with sodium-citrate as an anticoagulant [14].

DNA samples were used as templates in polymerase chain reaction (PCR). For each sample, a single PCR was performed using primers A, B and C. A 356 bp fragment was amplified by using primers A (5’-AATCATTCGTC TGTTTCCCA-3’) and B (5’-GTTTTCCTACCCTCAGA TT-3’). With these two primers only fusion db genes can be detected. A 751 bp fragment was amplified by using primers A and C (5’-CCAATCTACTCCCAG GAGCA-3’). This fragment can be obtained only in the presence of a normal b-globin gene.

Finally, the sequence of a 356 bp fragment was determined by the dideoxy chain termination method using fluorescently labeled dideoxy nts (Pharmacia Biotech, Uppsala, Sweden) on an automated DNA sequencer (ALFexpress DNA Sequencer; Pharmacia Biotech), using primers A (as forward) and B (as reverse).