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The breakpoints of the translocation t(2;5)(p23;q35) associated with Ki-1-positive anaplastic large-cell lymphoma (Ki-1 ALCL) involve a novel tyrosine kinase gene, ALK, at 2p23 and the nucleophosmin gene, NPM, at 5q35. Reverse transcriptase-polymerase chain reaction (RT-PCR) using NPM and ALK primers detects a consistent fusion product in Ki-1 ALCL cases with the translocation, resulting from genomic breakpoints within the same respective introns of NPM and ALK. To examine the feasibility of long-range DNA PCR with the same exonic NPM and ALK primers for the detection of the genomic NPM-ALK rearrangement, we examined 20 cases of Ki-1 ALCL previously characterized by NPM-ALK RT-PCR. Ten cases were positive for the NPM-ALK fusion RNA and 10 were negative. We first confirmed that both the NPM and ALK normal introns are relatively short, approximately 1 and 2 kb, respectively, suggesting that the largest possible size for the chimeric NPM-ALK intron would be about 3 kb. All 10 cases positive by RT-PCR were also positive by long-range DNA PCR. The DNA PCR products ranged, as expected, from the sizes of the normal introns, between 0.5 and 2.5 kb. All 10 RT-PCR-negative cases were also negative by long-range DNA PCR, and control templates for RT-PCR and long-range DNA PCR were successfully amplified. Thus, we have shown that the introns involved by the NPM-ALK rearrangement seen in some Ki-1 lymphomas are relatively short, making the genomic rearrangement amenable to reliable detection by longrange DNA PCR. Furthermore, the variability observed in the sizes of chimeric introns is evidence against clustering of the genomic breakpoints within these introns.