Study On The Polyc Ationic Gene Vector Which Was Synthesized From The Endogenous Substance

As a vector for gene delivery, cationic polymers possess several advantages, such as the safey, greater loading capacity and loading density, simple and controllable preparation, easier modification and more stable compared to the viral vectors and cationic liposomes. However, the cationic polymers also suffer from two major deficiencies: high cytotoxicity and low transfection efficiency. Up to now, there are several kinds of cationic polymers which all stay on the phase of experiment reagent. So, to design a successful vector, we should take into account a series of biological processes, including condensing DNA, circulating in vivo, targeting to special cells, escaping from lysosome, releasing DNA and degrading. According to this, we synthesized a novel polycation for gene delivery with a natural molecular existed in the human sperm.There are three parts in this dissertation. I introduced the polycation from three aspects: synthesize and chemical characterization, physical chemistry and bioactivity.First, we used ethylene bis-(chloroformate) to link spermine which had had their primary amine protected and got a linear polymer. Through the chemical characterization, we can confirm we have got what we want. The polymer’s molecular weight is 1144 and it has the longer half life of degradation than our anticipation.Then it is the physical chemistry characterization. The Agarose Gel Electrophoresis showed that the plasmid was fully protected at a ratio of 3:1 w/w. The zeta potential and particle size of the complexes were respectively 33mV and 50nm, fell into the suitable range. So we can get a compact and right size particle.To determine the bioactivity, COS-7 and 293ft cells lines are used to evaluate the transfection efficiencies and cytotoxicity. We found that polymer have high transfection efficiency to the COS-7 cells and 293ft cells, almost 2 times as that of PEI 25kDa, at the same time, it also had a certain activity of gene silencing by delivering siRNA to HSC. We use the taqman probe to track the polylink-SP by using laser confocal Microscope. We can confirm that the polymer can take the gene out of the lysosome successfully and then target to the surface of nucleus membrane. The transfection and cytotoxicity assay suggested that the polymer has the greater transfection efficiency but less toxicity compared with PEI25kDa, which would make it to be a potential nontoxic cationic polymeric gene carrier.To summarize, our research synthesize a novel polycation gene vector with a natural molecular in the human sperm. It can condense gene efficiently, escape from the lysosome and protect the gene against the degradation. Also, it has lower cytotoxicity and still has the space for improvement in its construct. It provides the new thought for further application with non-viral vector in clinic research.