GENE-TRANSFER INTO MOUSE LYOMA CELLS BY ELECTROPORATION IN HIGH ELECTRIC-FIELDS
GENE-TRANSFER INTO MOUSE LYOMA CELLS BY ELECTROPORATION IN HIGH ELECTRIC-FIELDS
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DOI：
10.1002/j.1460-2075.1982.tb01257.x
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发表时间：
1982-01-01
期刊：
影响因子：
11.4
通讯作者：
HOFSCHNEIDER, PH
中科院分区：
文献类型：
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作者：
NEUMANN, E;SCHAEFERRIDDER, M;HOFSCHNEIDER, PH
电脉冲（8 kV/cm，5 μ S）极大地增加了细胞对DNA的摄取。当含有单纯疱疹胸苷激酶（TK）基因的线性或环状质粒DNA加入到TK基因缺陷的小鼠L细胞悬浮液中，然后将细胞暴露于电场时，形成了在HAT [次黄嘌呤-氨基蝶呤-胸苷]选择培养基中存活的稳定转化体。在20度。在施加3个连续的电脉冲后10分钟使DNA进入，结果为95 ± 0.01 μ g/ml。3)每106个细胞和每1.2 μ g DNA的转化体数。与生物化学技术相比，电场法基因转移具有操作简单、易于实施、效率高等优点。由于DNA的细胞膜转运机制尚不清楚，本文提出了一个简单的物理模型，用于描述在高电场下DNA穿透细胞的增强过程。根据该电穿孔模型，外部电场与孔配置的脂质偶极子的相互作用诱导并稳定渗透位点，从而增强跨膜转运。
Electric impulses (8 kV/cm, 5 .mu.S) increased greatly the uptake of DNA into cells. When linear or circular plasmid DNA containing the herpes simplex thymidine kinase (TK) gene is added to a suspension of mouse L cells deficient in the TK gene and the cells are then exposed to electric fields, stable transformants are formed that survive in the HAT [hypoxanthine-aminopterin-thymidine] selection medium. At 20.degree. C after the application of 3 successive electric impulses followed by 10 min to allow DNA entry there resulted 95 (.+-. 3) transformants per 106 cells and per 1.2 .mu.g DNA. Compared with biochemical techniques, the electric field method of gene transfer is very simple, easily applicable, and very efficient. Because the mechanism of DNA transport through cell membranes is not known, a simple physical model for the enhanced DNA penetration into cells in high electic fields is proposed. According to this electroporation model the interaction of the external electric field with the lipid dipoles of a pore configuration induces and stabilizes the permeation sites and thus enhances cross membrane transport.