Electroporation-mediated gene transfer system applied to cultured CNS neurons
Electroporation-mediated gene transfer system applied to cultured CNS neurons
复制标题
DOI：
10.1097/00001756-200404290-00008
复制
发表时间：
2004-04-29
期刊：
影响因子：
1.7
通讯作者：
Okabe, S
中科院分区：
文献类型：
--
作者：
Kawabata, I;Umeda, T;Okabe, S
电穿孔可以有效地将外源基因转移到完整脑组织中未成熟的神经元中。我们利用这种方法将基因转染到发育中的啮齿动物海马体中。随后解离转染的海马并使其在培养物中分化。通过优化海马体的发育阶段、驱动标记cDNA的启动子以及培养条件，神经元在电穿孔后两周以上仍保持多个标记基因的强表达。我们还可以通过将含有loxP侧翼终止序列的质粒的电穿孔与产生Cre的重组腺病毒的感染相结合来诱导成熟神经元中的瞬时表达。这里描述的系统可用于分析多个基因在神经元发育特定阶段的生物学作用。
Electroporation is effective in transferring foreign genes into immature neurons in intact brain tissue. We utilized this approach to transfect genes into developing rodent hippocampi. Transfected hippocampi were subsequently dissociated and allowed to differentiate in culture. By optimizing developmental stage of the hippocampus, promoters to drive the marker cDNA, and culture conditions, neurons kept strong expression of multiple marker genes for more than two weeks after electroporation. We could also induce transient expression in mature neurons by combining electroporation of plasmids containing loxP-flanked stopper sequences and infection of Cre-producing recombinant adenoviruses. The system described here is useful in analyzing biological roles of multiple genes in specific stages of neuronal development.