Figure 3.

Intracellular localization of replicated plasmid containing tandem lacO repeats by staining with an MPB-lacI fusion protein. HeLa cells were transfected with pSV2-dhfr-8.32 containing lacO tandem repeats and 24 h later were infected with 3 PFU per cell of vV5D4 expressing V5-tagged uracil DNA glycosylase. At 12 h after infection, cells were fixed, permeabilized, incubated with MBP-lacI and rabbit antibody to MBP (anti-MBP) and mouse monoclonal antibody to the V5 epitope (anti-V5) followed by Cy5-conjugated donkey anti-mouse IgG and Texas red dye conjugated donkey anti-rabbit IgG. Cells were counterstained with Hoechst dye and analyzed by confocal microscopy. Colors: deep blue, Hoechst dye; red, Texas red; white, Cy5; light blue, overlap of Texas red and Cy5; yellow, overlap of Hoechst and Cy5.