Figure 4

Engineering and fluorescence microscopy of αDENV-GrpIs in a Bicistronic Plasmid. A) Each of the trans-splicing αDENV-GrpI was tagged downstream of the firefly (FL) 3' exon with the mCherry fluorescent marker gene expressed from an IRES sequence of either the Black Queen Cell Virus (BQCV) or Drosophila C Virus (DCV). Expression of these constructs was driven by the Drosophila melanogaster Actin 5c promoter. This bi-cistronic configuration allowed monitoring for the presence and expression of the αDENV-GrpI constructs within cell cultures. A5c = Drosophila melanogaster actin 5c promoter, IRES = DCV or BQCV Internal ribosome entry site. B) Expression of mCherry was verified for each construct by transfecting 1 ug of plasmid DNA into C6/36 cells and examining at 48 hours post transfection. Photographs were taken under 40× magnification.