DETECTION OF 4 DIFFERENT 11Q23 CHROMOSOMAL-ABNORMALITIES BY MULTIPLEX-PCR AND FLUORESCENCE-BASED AUTOMATIC DNA-FRAGMENT ANALYSIS

A nested polymerase chain reaction (PCR) protocol was developed for ra pid detection of four different 11q23 abnormalities by a single PCR as say. During each of the two PCR rounds a sense primer located within e xon 5 of the MLL gene at 11q23 was combined with four different antise nse primers, each located within possible translocation partner genes at chromosomes 4, 6, 9, and 19, respectively. Except for the MLL prime r all primers used during the second round of nested-PCR carried a cha racteristic fluorescence label at their 5'-end. Agarose gel analysis o f the PCR products was sufficient to discriminate between the absence of any of the four MLL rearrangements and the presence of at least one of them. Discrimination of the four different MLL translocation partn er genes was not possible by agarose gel analysis due to a molecular h eterogeneity of the 11q23 breakpoints resulting in PCR products of var iable size. For this reason, automatic fluorescence-based DNA-fragment analysis was used to exactly define the MLL translocation partner gen es if a positive result had been obtained by agarose gel analysis. In patients with leukemia, this assay may enable a fast and highly sensit ive detection of different 11q23 abnormalities, which usually correlat e with poor clinical prognosis.