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Author:
Date:
2018-03-20
[Abstract] This protocol describes the technique of ex-vivo electroporation to target embryonic hippocampal progenitors in an organotypic slice preparation. This technique allows gene perturbation for examining developmental processes in the embryonic hippocampus while retaining the environment and connectivity of the cells. Gene perturbation can include Cre-mediated recombination, RNAi-mediated knockdown, gene overexpression, or a combination of any of these. Ex-vivo electroporation can be performed at a wide range of embryonic stages, giving temporal control to the experimenter. Spatial control can be achieved more easily by preparing the brain in a Petri dish to target particular regions of the hippocampus. The electroporated explant cultures provide a highly tractable system ...
[摘要] 该协议描述了在器官切片制备中靶向胚胎海马祖细胞的体外电穿孔技术。 该技术允许基因摄动检查胚胎海马体的发育过程，同时保持细胞的环境和连接性。 基因扰动可以包括Cre介导的重组，RNAi介导的敲低，基因过表达或这些中任何的组合。 可以在广泛的胚胎阶段进行体外电穿孔，为实验者提供时间控制。 空间控制可以通过在皮氏培养皿中准备大脑来瞄准海马的特定区域来实现。 电穿孔的外植体培养物提供了用于研究包括祖细胞增殖，迁移和细胞命运获取在内的发育过程的高度易处理的系统。
【背景】由于其位于尾端的端脑，海马在可达性方面提出了挑战。胚胎海马甚至更难以接近，需要用子宫外科手术方法进行实验操作。器官切片文化规避了这个问题，同时保留了海马场细胞构筑学的许多方面，包括分子特征和连接性。尽管有描述来自啮齿类动物大脑的海马外植体产后培养的规程（Stoppini等人，1991; ...
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