Saito Tetsuichiro
Department of Developmental Biology, Graduate School of Medicine, Chiba University, Chiba 260-8670, Japan.
Nat Protoc. 2006;1(3):1552-8. doi: 10.1038/nprot.2006.276.
This protocol describes a basic method for in vivo electroporation in the nervous system of embryonic mice. Delivery of electric pulses following microinjection of DNA into the brain ventricle or the spinal cord central canal enables efficient transfection of genes into the nervous system. Transfection is facilitated by forceps-type electrodes, which hold the uterus and/or the yolk sac containing the embryo. More than ten embryos in a single pregnant mouse can be operated on within 30 min. More than 90% of operated embryos survive and more than 90% of these survivors express the transfected genes appropriately. Gene expression in neurons persists for a long time, even at postnatal stages, after electroporation. Thus, this method could be used to analyze roles of genes not only in embryonic development but also in higher order function of the nervous system, such as learning.
本方案描述了一种在胚胎小鼠神经系统中进行体内电穿孔的基本方法。在将DNA微量注射到脑室或脊髓中央管后施加电脉冲，可使基因高效转染到神经系统中。镊子型电极有助于转染，该电极可夹住含有胚胎的子宫和/或卵黄囊。在30分钟内可对一只怀孕小鼠体内的十多个胚胎进行操作。超过90%的手术胚胎存活，其中超过90%的存活胚胎能正确表达转染基因。电穿孔后，神经元中的基因表达会长期持续，甚至在出生后阶段也是如此。因此，该方法不仅可用于分析基因在胚胎发育中的作用，还可用于分析基因在神经系统高级功能（如学习）中的作用。
