Abstract:

Objective To optimize the complexation condition of the novel nano non-viral gene vectors of polycaprolactone (PCL)-polyethylene glycol (PEG)-chitosan with plasmid in vitro, and evaluate the transfection activity of the complexes. Methods The minimum N/P ratio of the complexation of non-viral gene vector with plasmid carrying green fluorescent protein gene (pEGFP) was measured by gel electrophoresis in vitro. The particle sizes of the complexes under different N/P ratios and different buffering conditions were determined by dynamic light scattering method. The 293T cells were transfected with complexes of pEGFP with PCL-PEG-chitosan, and the transfection efficiency of complexes was observed under fluorescence microscope. Results The results of gel electrophoresis demonstrated that the minimum N/P ratio of complexation of PCL-PEG-chitosan with pEGFP was 4∶1, the minimum N/P ratio of complexation of PCL-chitosan with pEGFP was also 4∶1, and the minimum N/P ratio of complexation of chitosan with pEGFP was 1∶1. Dynamic light scattering test revealed that the particle sizes of the complexes of PCL-PEG-chitosan with pEGFP decreased with the increase of N/P ratios, and the particle sizes in acetate buffer system and DMEM buffer system were similar and maintained stable. Fluorescence microscopic observation indicated that the transfection efficiency of PCL-PEG-chitosan improved to some extent. Conclusion PCL modification can attenuate the complexation ability of chitosan with pEGFP, and increase the minimum complexation N/P ratio. The transfection efficiency of PCL-PEG-chitosan improves after PCL and PEG modification, and needs to be further improved.

Key words: PCL-PEG-chitosan, modification, DNA, gene transfection, N/P ratio