Fig. 2

Southern blot of subcloned cell lines expressing various levels of beta-lactamase reporter reveals their genetic similarity. HeLa cells were transfected with pDES46. To evaluate reporter expression, they were stained with CCF2-AM and examined by fluorescence microscopy. Individual, subcloned cells were derived by limiting dilution. For Southern blots, DNA was extracted, 10 mcg was restricted with EcoRI, electrophoresed on a gel, blotted, and probed for TEM1 reporter. Reporter expression phenotypes: lanes 1–6, clones 1 and 2 and their derived subclones, variegating phenotype with mixed beta-lactamase (TEM1) expression; lanes 7–8, clone 3 and its derived subclone, predominantly blue cells with high levels of beta-lactamase (TEM1) expression. Cell line names: Clone 1: cell line 5B 0.3c/w C9 (parent of sub-clones corresponding to lanes 2 and 3)‬‬‬‬‬‬; subclone 1.1: 5B #1G10; subclone 1.2: ‬‬‬‬‬‬5B #4D9‬‬‬‬‬‬; clone 2: 6I 0.3c/w C8 (parent of sub-clones corresponding to lanes 5 and 6)‬‬‬‬‬‬; subclone 2.1: 6I #8G2‬‬‬‬‬‬; subclone 2.2: 6I #9 F8‬‬‬‬‬‬; clone 3: 1 1c/w B11 (parent of sub-clone in lane 8)‬‬‬‬‬‬; subclone 3.1: 1 #5H5‬‬‬‬‬‬. Control samples: lanes 9–10, negative, untransfected HeLa cells; and lanes 11–12, pDES46 plasmid DNA (50 and 500 pg). White arrows: TEM1 bands shared between cell clones and pDES46 L1 donor plasmid; black arrows, integrated TEM1 bands present in all clones, but not in EcoRI- digested pDES46 ‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬