载基因 PGenesil-TGF-β1壳聚糖纳米粒的制备和性能研究

摘要

To prepare chitosan nanoparticles carrying recombinant plasmid and investigate their performance.The chitosan -PGen-esil -TGF -β1 (reporter gene)nanoparticles were prepared by a complex coacervation process.The morphology and particle diameter of nanoparticles were observed under transmission electron microscope;the encapsulating rates were determined using full spectrum spectrophotometer;the binding of PGenesil -TGF -β1 to chitosan nanoparticles was evaluated by agarose gel electrophoresis analysis and the protective effect of nanoparticles against Deoxyribonuclease was analyzed by DNase I digestion experiment .The prepared chi-tosan -PGenesil -TGF -β1 nanoparticles were mainly spherical and the diameter distribution was 100 ~200 nm,with an average di-ameter of 127.37 ±19.75 nm;the encapsulation rate of chitosan -PGenesil -TGF -β1 nanoparticles was (94.38 ±0.45)%;the agarose gel electrophoresis analysis confirmed that the nanoparticles could combined plasmids through electrostatic interactions ,and DNase I digestion experiment demonstrated the nanoparticles could protect plasmids effectively from nuclease degradation.Chitosan nan-oparticles are successfully prepared by complex coacervation,and it will lay the foundation for further research on gene treatment.%制备负载重组质粒 PGenesil-TGF-β1的壳聚糖纳米粒，并研究其结构特征和性能特点。采用复凝聚法制备壳聚糖-PGenesil-TGF-β1（报告基因）纳米微粒体，通过投射电镜测定其形态、粒径；采用全光谱分光光度计测定该复合物的包封率；凝胶电泳阻滞实验分析纳米粒载体与质粒的结合能力；DNase I 消化实验观察壳聚糖纳米粒保护质粒抵抗核酸酶的能力。制备的壳聚糖-PGenesil -TGF -β1纳米粒均呈球形，微粒直径在100～200 nm 之间，平均约（127．37±19．75）nm；其包封率为（94．38±0．45）％；凝胶电泳阻滞实验结果表明壳聚糖纳米粒能通过静电作用有效结合质粒，将其包裹在内；DNase I 消化实验显示壳聚糖纳米粒能有效地保护质粒免受核酸酶的降解。用复凝聚法成功制得壳聚糖-PGenesil -TGF -β1纳米粒，为后续研究基因药物奠定了实验基础。