SINCMV vector design and transcript expression in retroviral producer cells. A. The control SIN vector lacking an internal promoter has major deletions in the 3'LTR enhancer elements rendering its intrinsic promoter machinery transcriptionally inactive in transduced cells. B. Transgene expression in the SINCMV design is driven by the internal CMVIE promoter embedded upstream of the reporter EGFP. Three RNA transcripts are expected from SINCMV proviral DNA transcription in transfected packaging cells. The upstream CMV promoter in the 5'LTR drives the expression of a full-length ~2.6 kb transcript that can be packaged into retroparticles and a ~2.1 kb spliced form that lacks the packaging signal (Ψ). The internal CMVIE drives the expression of a shorter ~1 kb transcript. C. Hybridization with a P32- labelled EGFP probe done on total RNA extracted from SINCMV retroviral producers treated with butyrate indicated significant increase in the level of retrovector mRNA. D. Loading control of the 3 RNA samples is shown by ribosomal RNA staining with ethidium bromide.