Effect of Sam68ΔC on association of env RNA with polysomes. Cells were co- transfected with Rev, the HIV-1 env expression plasmid pgTat and pcDNA or Sam68ΔC. 48 hours post-transfection cells were harvested and cytoplasmic extracts layered onto 15–50% linear sucrose gradients. Following centrifugation, gradients were fractionated and analyzed by monitoring (a) absorbance at 254 nm (b) distribution of Sam68ΔC and ribosomal protein L26 by western blotting or (c) actin and (d) unspliced env RNA distribution by QRT-PCR. To ascertain whether profiles were dependent upon the integrity of polysomes, an aliquot of the cell lysate was treated with EDTA (+EDTA) to dissociate ribosomal subunits prior to fractionation on the gradients.