Figure 1

Co-ordinated co-expression to different compartments in HeLa cells. A single ORF was designed with the fluorescent genes eyfp and ecfp plus the puromycin resistant gene pac [9]. These genes were cloned flanking FMDV 2A sequences. An internal signal-anchor from the human β-1,4 galactosyltransferase (GT) was fused to the 5' end of the ecfp for Golgi targeting. During its translation, the self-processing of this polyprotein produced EYFP-2A that diffused to the cytoplasm and nucleus (due to its small size), while GT-EYFP-2A was co-translationally targeted to the Golgi apparatus by the GT signal (some protein also stays in the endoplasmic reticulum, due to the continuous cycling between these compartments). Two fields are shown, in both cases the cell on the left shows a high level of expression of both proteins that were expressed at lower levels in the cell on the right, illustrating the co-ordination obtained with the 2A co-expression strategy. PAC was able to confer resistance to puromycin. Images were taken 48 hours post-transfection. Bar represents 10 μm.