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. Author manuscript; available in PMC: 2010 Aug 1.
Published in final edited form as: Gene Ther. 2009 Oct 22;17(2):217–226. doi: 10.1038/gt.2009.136

Figure 2. Effect of nuclear localization of integrase on transgene expression in cell culture and in liver.

Figure 2

(a) HeLa cells were transfected using FuGene 6 in a 1:50 ratio of donor plasmid (pDB2) to integrase plasmid (pCSmI, inactive integrase, green; pCSI, wild-type integrase, pink; pCSI-HA, wild-type integrase with HA tag, orange; pCSI-HA-NLS, wild-type integrase with NLS after HA tag, blue; pCSI-NLS-HA, wild-type integrase with NLS before HA tag, red). Stable integrants were selected with G418, and the colonies that formed after two weeks that were greater than 1 mm in diameter were counted. p<10-7 for two-tailed t-test comparing pCSI-HA to pCSI-HA-NLS or pCSI-NLS-HA. (b) A solution containing 20 μg each of donor plasmid (pVFB, hAAT liver-specific promoter driving human Factor IX with an attB) and integrase plasmid (pCSmI, green diamond; pCSI, pink square; pCSI-HA-NLS, blue circle; pCSI-NLS-HA, red triangle) was administered to 8-10 week old female C57BL/6 mice by hydrodynamic injection. Mice were bled at the indicated time points, and ELISAs were done to determine hFIX levels in the serum. Control data were in common with another, simultaneous study, to reduce animal usage.35 (c) Staining of liver sections from mice in (b) for hFIX (green). Error bars are the standard error of the mean in (a) and (b); n ≥ 5 in (a) and (b). *p<0.005; two-tailed t-test performed on end-point data for each group, pCSI, pCSI-NLS-HA, or pCSI-HA-NLS against the pCSmI group.