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. 2005 Apr 15;6(5):458–463. doi: 10.1038/sj.embor.7400390

Figure 1.

Figure 1

Construction and transfection of plasmids in C2C12 myoblast cells. (A) Plasmids containing the wild type (UTR(wt)), mutant (UTR(200)), mutant with the WPRE sequence or its antisense version (UTR(200W) and UTR(200W-AS), respectively) were constructed. All plasmids coexpressed the EGFP gene under the Rosa26 promoter. Arrows indicate binding sites for PCR primers used in quantitative RT–PCR assays in Fig 3. (B) After transient transfections of the above constructs in C2C12 cells, low levels of EGFP were observed in cells transfected with the UTR(200) construct, whereas the addition of the WPRE sequence caused a significant increase in the EGFP expression (UTR(200W)). Normal EGFP levels were obtained with the UTR(wt) construct, whereas no EGFP was seen in untransfected cells (UN). The effect of WPRE was specific as no increase in EGFP levels was seen in cells transfected with its antisense version (UTR(200W-AS)). Nuclear staining by 4,6-diamidino-2-phenylindole (DAPI) shows the presence of cells. Scale bar, 0.1 mm. (C) Western blot analysis against EGFP showed results similar to (B), indicating that addition of the WPRE sequence increases transgene expression.