Figure 3
From: Characterization of a potent non-cytotoxic shRNA directed to the HIV-1 co-receptor CCR5
Efficient reduction requires short hairpin structure and the mode of action is consistent with RNAi. huCCR5-293T cells (0.5 × 105) were plated into 24 well plates one day before infection. Cells were transduced with lentiviral vectors for 2 hrs in the presence of 8 μg/mL polybrene. The transduced cells were harvested 3 days later and stained with APC conjugated anti-human CCR5 monoclonal antibody for flow cytometry. The efficiency of CCR5 reduction was compared in EGFP+ cells transduced by lentiviral vectors expressing no shRNA (negative control), CCR5 shRNA driven by the U6 {U6-shRNA (1005)} or the H1 promoter {H1-shRNA (1005)}, sense and antisense siRNA expressed from independent U6 promoters from a vector (U6-sense U6-antisense siRNA) or (U6-antisense siRNA). The x axis indicates EGFP expression; the y axis indicates CCR5 expression. The percentage of cells in each quadrant is shown. The quadrant lines were defined by mock-transduction cells. Mock: uninfected cell.