CXCR4/MAML2 fusion gene is expressed in CLL. (A) Top panel, EcoRI restriction map around the breakpoint sites on chromosomes 2 and 11. The green line represents the full-length EcoRI-digested fragment without any break; the light blue line represents the fragment length after chromosome break and the EcoRI-digested fragment after CXCR4/MAML2 fusion. Bottom panel, Southern Blot EcoRI digested. The CXCR4/MAML2 gene fusion fragment of ∼7 kb is shown on lane 2 (GI-403 clone) and on lane 4 (patient B cells). The probe has been designed on the MAML2 gene and its amplification primers are reported in supplemental Table 2. (B) PCR on total cDNA performed with “fusFW” and “fusRV” oligos, designed to amplify a region of 330 bp across the exon1 CXCR4 and the exon2 of MAML2. Primer sequences are reported in supplemental Table 2. The amplicons obtained from this PCR were sequenced (reverse primer used). (C) Western blot using anti-MAML2 polyclonal antibody. The chimeric protein CXCR4/MAML2 (lanes 3 and 5) has a smaller size (∼20 kDa) than MAML2 wt. (D) Ponceau staining of the nitrocellulose membrane shows the total proteins are equally loaded in each lane. This control is relative to panel C.