Correlation of eGFP fluorescence and mRNA transgene expression levels. Four independently produced preparations of CH-eGFP-WS-derived lentiviral vector were serially diluted (1/10) prior to transduction of 293T cells. 6 days later, cells were harvested for eGFP analysis by FACS or RNA extraction and subsequent RT-qPCR with primers and probe directed against WPRE to measure transgene expression and against RNAse P to normalize for total RNA content. Total eGFP expression was measured by multiplying the percentage of transduced cells (TE) with the mean fluorescence intensity (MFI). The transgene mRNA level is given as the number of mRNA copies normalized to the total RNA content (RNAc/ng). These values correlate strongly with each other (r2 = 0.97).