IPCR analysis to detect etoposide-induced rearrangements initiating within the MLL bcr. (A) Schematic representation of the MLL gene locus on chromosome band 11q23. The 8.3-kb MLL bcr, flanked by BamHI sites, includes exons 5 to 11 and intervening introns. XbaI sites are 2.6 kb apart within the MLL bcr. Genomic DNA was digested with XbaI and circularized. Nested PCR reactions were carried out with F1-R1 primer pair, followed by F2-R2 primer pair. BH (BamH1); X (XbaI); F1, F2, R1, R2 are primers used for PCR (see “Materials and methods”). (B) Representative IPCR products. Lanes 1-7: untreated controls that give the expected 1.8-kb germ-line product. Lanes 8-18: etoposide-treated samples that all give alternative products representing possible rearrangements of MLL. Pretreatment of samples shown in lanes 8 to 18 with PvuII eliminated some or all detectable germ-line product to facilitate isolation of individual alternative sized products. Lanes 8-12: multiple independent CD34⁺ cell samples (long recovery) with all germ-line products eliminated by PvuII in lanes 8-9; lane 13: TF-1 (short recovery); lane 14: WS1 (short recovery); lanes 15-16: M059K (short recovery); lane 17: M059J (short recovery); and lane 18: cord blood mononuclear cells (short recovery). Parallel experiments in multiple hematologic and other adherent cell lines gave similar IPCR results (B), but individual repair clones have not been fully characterized.