The humanMLLgene is one of the most promiscuous recombination hot spots of our genome with regard to the onset of malignant diseases. With the exception of gene internal partial-tandem duplications involving several exons located in the 5′-end ofMLL, all recombination events occur in a small genomic region flanked byMLLexons 8–14, designated as theMLLbreakpoint cluster region. Efforts from different laboratories, including our own, have led to the identification of more than 50MLLfusion partners that were characterized at the molecular level. The common theme of recombination events involving the humanMLLgene is the creation of “functional” fusion genes that are translated into oncoproteins. Many different labs have already demonstrated that these MLL fusion proteins have the capability to instruct hematopoietic stem/precursor cells to convert into a preleukemic state, which finally leads to the onset of leukemia in experimental model systems. Here we have focused on the identification ofMLLfusion partners by using the genomic DNA of acute leukemia patients. After initial screening using for example split signal FISH experiments (as an example for technologies to identifyMLLrearrangements), genomic DNA from leukemia patients is analyzed by long-distance-inverse (LDI)-PCR. LDI-PCR is based on the hydrolysis of patient DNA using distinct combinations of restriction enzymes, self-ligation of the resulting DNA fragments and a subsequent PCR reaction using a specific set of oligonucleotides. This strategy allows in principle any investigator to identify known and unknownMLLfusion partner genes. Furthermore, the genomic fusion site inMLLrearrangements represent a unique and reliable molecular marker that allows the tracing of minimal residual disease (MRD) in these patients before, during, and after therapy.
