Retroviral vectors and experimental design. (A) The HOXB6 vector was constructed by cloning the cDNA encoding HOXB6, modified by the addition of an N-terminal FLAG epitope into the multiple cloning site of the MSCV-IRES-EGFP control vector. The HOXB6WG vector was constructed by introducing a single amino acid W → G substitution at position 130 within the conserved YPWM PBX-interaction motif (PIM). Similarly, the HOXB6NA vector includes one amino acid N → A substitution at position 196 within the homeodomain. The unique XhoI and ClaI restriction sights used for Southern blot analysis are shown. Digestion with both enzymes releases a 1.3-kb proviral fragment, whereas digestion with BamHI or XhoI alone cuts the provirus once, thus generating unique-sized fragments for individual viral integrations. LTR indicates long terminal repeat; F, N-terminal flag epitope; HD, homeodomain; 5FU, 5-fluorouracil; and RV, retroviral. (B) Schematic representation of experimental design.