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. 2013 Dec 9;203(5):767–783. doi: 10.1083/jcb.201305027

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© 2013 Bian et al.

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Figure 1. — 200-kb human β-globin BAC transgene targets to nuclear periphery similarly to endogenous β-globin locus. (a–c) FISH shows peripheral localization of endogenous β-globin locus in human BJ-hTERT (a) and mouse NIH 3T3 (b) fibroblasts but interior localization for the α-globin locus in mouse NIH 3T3 cells (c; DAPI [blue], FISH [green], and lamin A immunostaining [red]). Arrowheads show DNA FISH signals. (d–f) Peripheral localization of HBB BAC identified by EGFP-LacI binding (green) in HBB C3 NIH 3T3 cell clone using DAPI staining (blue; d), lamin A immunostaining (red; e), or nuclear pore staining (red; f) to define the nuclear periphery. Arrowheads show the HBB transgenes. (g) Fraction of cells with peripheral localization in each NIH 3T3 subclone for HBB (black) or DHFR (green) BAC transgenes as compared with endogenous β-globin loci (HBB) in human BJ-hTERT (dark gray) or mouse NIH 3T3 (red) cells or α-globin loci (HBA) in human BJ-hTERT cells. HBB BAC (no LacO; yellow) refers to FISH measurements from a mixed population of stable NIH 3T3 clones with HBB BAC with just a selectable marker but no LacO repeat inserted. Random shows a fraction of DAPI staining within 0.5 µm from the periphery. At least 50 cells from each BAC transgene NIH 3T3 cell clone and ≥45 cells for each endogenous gene FISH experiment were analyzed. Bars, 2 µm.