High-sensitivity analysis of the 1355insAA-duplication nt 1324-1355 before and after BMT. (A) Schematic representation of the insertion-duplication in exon 6 of the PIG-A gene. At the top, two copies of the normal sequence are aligned to show direct-sequence homology (underlined) that may favor nonhomologous exchange below the sequence in the recent sample of patient MSK13. Identical bases are indicated by a vertical line. The short repeat of 4 nt flanking the duplication are boldface. The 19 mer D has been designed to match completely the mutant sequence, whereas the last 4 nt (5′-… .ttTC-3′) are mismatched to the normal sequence (see arrow bent at the left). The fragment amplified with primers -I5b and D and -I5b and f are expected to be 292 bp and 415 bp in length, respectively. (B) Gel electrophoretic analysis of nested-PCR products of exon 6 of PIG-A gene from PMN in the pretransplant sample. A fragment of 292 bp amplified with the primers -I5b and D was present in the post-transplant sample (lane 2); by contrast no amplification was obtained in the pretransplant sample (lane 1) and in a normal control sample (lane 3). When the 3 primers -I5b, D, and f were used in the same PCR reaction, a fragment of 415 bp only was amplified in the pretransplant sample and in a normal control (lanes 4 and 6), and a fragment of 292 bp only was amplified in the posttransplant sample (lane 5).