Abstract:

 Objective     To investigate the effect of a new cationic carrier polyethylenimine (PEI) on transfection of the aquaporin 5 (AQP5) gene into A549 cells in vitro. Methods      According to different N/P ratios (5, 10, 15, 20 and 25), the eukaryotic expression vector pEGFP-N1 was mixed with the green fluorescent protein(GFP) and PEI to make a transfection complex, using lipofectamine2000 as a positive control. Transfection efficiency was detected by flow cytometry for optimizing the optimal N/P ratio.  Fulllength AQP5 cDNA were cloned into the pEGFP-N1 vector to construct pEGFP-N-AQP5, which was then transfected into A549 cells. Expression of AQP5 in transfected cells was detected using PCR and Western blot. Results      Flow cytometry revealed the GFP emitting. When the N/P ratio was 15, the PEI/pEGFP-N1 complex achieved the highest transfection efficiency of 89.72%, close to 91.44% when delivered by liposomes. Compared with the control group, AQP5 gene and protein expressions were significantly increased after successful transfection of pEGFP-N-AQP5 into A549 by application of the PEI and lipofectamine2000 complex. Conclusion      pEGFP-N-AQP5 can be successfully transfected into A549 cells mediated by the novel cationic carrier PEI.

Key words: Polyethylenimine; Transfection; Aquaprins