Figure 2 | Oncogene

Figure 2

From: Use of adeno-associated viral vector for delivery of small interfering RNA

Figure 2

Downregulation of p53 expression by AAV-delivered siRNA. (a) High-efficiency gene transduction by si-p53 encoding AAV vectors. HEK293 cells were cotransfected with pH1-si-p53/AAV-GFP, pAAV-RC and pHelper plasmids and recombinant adenoviral particles collected 3 days post-transfection according to the instructions of the manufacturer. Virus stock was frozen at −80°C prior to use. HeLa S3 cells were plated at a density of 1.5 × 105 cells in each well of a 12-well plate and next day infected with 0.5 ml of virus-containing supernatant. At 72 h postinfection, cells were examined under a fluorescent microscope to visualize the expression of GFP and photographed. (b) Suppression of p53 expression in HeLa S3 cells by AAV-delivered siRNA. HeLa S3 cells were infected with pH1-si-cont/AAV-GFP and pH1-si-p53/AAV-GFP viruses, respectively. Cellular lysates were prepared 48 h after infection. Western blot analysis demonstrates significant downregulation of p53 expression in pH1-si-p53/AAV-GFP-infected cells. Blot was reprobed with an antibody against actin to show equal loading of all lanes and to serve as a specificity control. Primary antibodies against p53 (clone DO-1) and actin were obtained from Santa Cruz Biotechnology (Santa Cruz, CA, USA)

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