脂质体介导的外源基因在黄瓜悬浮细胞原生质体中的表达

本文用脂质体包裹质粒DNA,研究了不同制备方法,不同磷脂成分对包裹率的影响,以及超声处理对脂质体内质粒DNA稳定性的影响。用改进的脱水再加水法制备质粒脂质体,使包襄率由10—20％提高到60—70％。黄瓜悬浮细胞原生质体与包裹pUC12—CAT质粒的脂质体经PEG诱导融合,培养3天和6天后均测得较强的CAT活性,说明脂质体介导的外源基因己在黄瓜细胞中成功地表达。

LIPOSOME-MEDIATED GENE TRANSFER AND TRANSIENT EXPRESSION IN CUCUMBER SUSPENSION PROTOPLASTS

The effects of different liposome preparation methods and the phospho-lipid composition on encapsulation efficiency of plasmid DNA were investigated. By a modified dehydration-rehydration method, the encapsulation efficiency was increased from 10-20% to 60-70%. Soybean phospholipid as a source for liposome preparation was found to be superior to either PC : chol:OA (4:4:3) or PE :chol:OA(4 :4: 3). It was evident that even short period of sonication (10-20 sec.) had a strong effect on the stability of plsamid DNA which resulted in a dramatic DNA degradation. Protoplasts isolated from cucumber suspension culture were fused by polyethylene glycol with liposomes in which a pUCl2-GAT ( CaMV 35S-CAT) gene was encapsulated. After 3 and 6 days in culture, the cells showed strong CAT activity indicating foreign gene was successfully expressed in cucumber cells.