Figure 4
HIV-1 Rev/RRE and cis elements in the 5'UTR do not influence vector titers after packaging into MLV viral particles. A. Titers of MLV/HIV chimeric vectors were obtained by scoring for GFP positive cells following transduction of 293T cells. Titers are expressed as transducing units (TU) normalized to the amount of RT units (counts per minute [CPM]). B. Normalized luciferase levels were determined in transfected 293T producer cells. Luciferase levels were normalized to total cell protein. C. Titers (part A) expressed as a ratio to levels of luciferase (part B) shown in arbitrary units (AU). All experiments were executed in the absence (white bars) and presence (black bars) of Rev. Error for all bar graphs is expressed as ±S.D. All experiments were performed in triplicate. D and E. 293T cells were transduced with equivalent amounts of RT units (6 × 105 CPM), as determined for each of the indicated chimeric vectors. The influence of the HIV-1 Rev/RRE system, and 5' UTR cis elements, on transduction was assessed by fluorescence microscopy D and FACscan analysis E at 7 days post-transduction. The percent GFP positive cells are indicated for each FACscan.