Transfection efficiency of various pc3.5-based vectors harboring promoters and/or introns controlling the gene of interest. Plasmids pEF3, pCOX2.5, pPGK1.5puro, pPGKi.5puro, pEF3.5bpuro, pCMVi.5puro, and pmaxCDK harboring the bicistronic insert EGFPEMCVChFP were expressed in (left) 293T cells, (middle) B16 cells and (right) mesenchymal stem cells (MSCs). These harbor the following promoters and/or introns: cyclooxygenase (COX)-2, phosphoglycerate kinase (PGK)-1, PGK-1 with β-globin/IgG chimeric intron, EF3.5b, cytomegalovirus (CMV) with β-globin/IgG chimeric intron, and CMV with β-globin/IgG chimeric intron. The asterisk implies expression of the insert from plasmid pmaxCDK instead of from plasmid pCMVi.5puro. Black bars, relative transfection efficiency compared with cells transfected with pEF3-EGFPEMCVChFP; grey bars, relative EGFP levels compared with cells transfected with pEF3-EGFPEMCVChFP. Plasmid pmaxGFP was also transfected to determine whether improved transfection in MSCs was obtained. Expression from the COX-2 promoter was insignificant and therefore not shown in B16 cells and in MSCs.