Tat activates translation of its cognate mRNA. Panel A shows the monocistronic plasmids encoding Rluc used in these experiments. Panels in B report the results obtained with increasing amounts of pTat DNA, from 0 to 20 ng (at least 3 independent assays were performed). In independent assays, optimal stimulation of Rluc expression was found to occur at 20 ng of pTat (Additional file 2). All results are reported as Rluc activity per RNA copy number monitored by RTqPCR (see methods). Note that the 5' UTR of Tat RNA is more active than that of the genomic RNA with or without pTat addition, in HeLa cells. Panels in C show that Tat did not influence expression of Rluc from plasmid pRenilla (top panel). In addition, the 5' TAR-pA sequences of the 5' UTR of Tat1 or Tat2 appear to be indispensable for Tat-mediated translational activation (lower panels).