摘要
为得到低毒、高效的聚阳离子基因载体,以甲基丙烯酸氨乙酯（AMA）和甲基丙烯酸N,N-二乙基氨乙基酯（DEAEMA）为单体,以2-溴代异丁酸乙酯（EBIB）为引发剂,通过原子转移自由基聚合（ATRP）制备了两种聚（甲基丙烯酸氨乙酯-co-甲基丙烯酸N,N-二乙基氨乙基酯）阳离子无规共聚物（P（AMA-co-DEAEMA）,简称P）.琼脂糖凝胶电泳实验结果表明聚合物P作为阳离子载体可以有效地络合DNA,通过粒径仪测定的复合物粒子的尺寸在400~600 nm之间.扫描电镜观察的P/DNA复合物形貌是分散均匀的球形颗粒.以25k Da PEI为阳性参照,利用MTT比色法考察了聚合物P对HEK293T细胞的毒性.结果表明,聚合物P的细胞毒性低于25 k Da PEI的细胞毒性.以25 k Da PEI和裸质粒DNA作为参照,我们进一步考察了聚合物P与DNA形成的复合物在HEK293T细胞中的转染效率.结果表明P/DNA复合物在HEK293T细胞中的转染效率远远高于裸质粒DNA的转染效率,并且接近于25 k Da PEI/DNA复合物的转染效率.
To develop low toxic and highly efficient gene vector, a series of random copolymers, poly （2- aminoethyl methacrylate-co-2-（ diethylamino ） ethyl methacrylate ） s （ P （ AMA-co-DEAEMA ） s, P）, were synthesized through atom transfer radical polymerization （ATRP）. Physiochemical characteristics of P/DNA complexes were analyzed by agarose gel electrophoresis, particle size measurement, and scanning electron microscopy （SEM）. The results of agarose gel electrophoresis retardation assay demonstrated that random copolymer P can effectively bind DNA. The particle size of P/DNA complexes was around 400 - 600 nm. SEM images showed that complexes displayed a regular spherical shape. In vitro cytotoxicity measurements showed that polymer P had much lower cytotoxicity than that of 25 kDa PEI. The transfection efficiency was evaluated in HEK 293T cells, and 25 kDa PEI as well as plasmid DNA were used as positive and negative controls, respectively. The transfection efficiency of P/DNA complexes at each N/P ratio was much higher than that of plasmid DNA,and comparable to that of 25 kDa PEI/DNA complexes at N/P ratio of 10. The much reduced eytotoxieity and high transfection efficiency indicate that polymer P will find great potential as a non-viral gene vector.
出处
《高分子学报》
SCIE
CAS
CSCD
北大核心
2015年第1期113-119,共7页
Acta Polymerica Sinica
基金
国家自然科学基金(基金号21004046)资助项目
关键词
基因载体
转染效率
细胞毒性
原子转移自由基聚合
Gene vector, Transfection efficiency, Cytotoxicity, Atom transfer radical polymerization
