Cord blood–derived IGF-2 RT-PCR products digested with ApaI andAvaII.

When digested with ApaI (A) or AvaII (B) they show monoallelic expression in specimens X254 and X256 but partial biallelic expression in X263 (arrows). Sample X263 also shows some shorter nonspecific (ns) PCR products that migrated in a similar position (below the bands of interest) before enzyme digestion. The ratio of the digested band intensity reflects the effect of heteroduplex formation on the digestion of PCR products. Results are consistent with a starting mix of A and B alleles of approximately 0.8:0.2. Assuming complete heteroduplex formation at the end of PCR cycling, the PCR products consist of a mixture of AA, AB, and BB duplexes in a ratio of a2:2ab:b2—ie, 0.64:0.32:0.04. On incubation with ApaI, only the BB homodimers are digested, resulting in a band ratio of 0.96:0.04 = 24:1 (undigested:digested), whereas incubation with the reciprocal enzyme AvaII products will only digest AA homodimers, resulting in a 0.36:0.64 ratio = 0.56:1. These ratios (24:1 and 0.56:1) are similar to those seen in Figure 5A and B, respectively.