Mauti Olivier, Baeriswyl Thomas, Stoeckli Esther T
Institute of Zoology, University of Zurich, 8057 Zurich, Switzerland.
CSH Protoc. 2008 Dec 1;2008:pdb.prot5094. doi: 10.1101/pdb.prot5094.
INTRODUCTIONIn ovo RNA interference (RNAi) is a method for silencing a gene of interest using a combination of in ovo injection and electroporation in avian embryos. Here we describe gene silencing in the developing spinal cord, but the procedure can easily be adapted to other parts of the nervous system. Double-stranded RNA (dsRNA) derived from the gene of interest is injected into the developing spinal cord of the chicken embryo, and is followed by electroporation to allow for the uptake of the dsRNA. With this method, temporal as well as spatial control of gene silencing is possible. The time point of injection should be chosen according to the expression profile of the gene or the half-life of the protein. Proteins with slow turnover may require RNAi at earlier stages, ideally before the onset of gene expression. The electroporation parameters can be adjusted such that only a specific population of neurons is targeted in the spinal cord.
引言
卵内RNA干扰（RNAi）是一种利用卵内注射和电穿孔相结合的方法，在禽类胚胎中沉默目标基因的技术。在此，我们描述了在发育中的脊髓中进行基因沉默的方法，但该程序可轻松适用于神经系统的其他部位。将源自目标基因的双链RNA（dsRNA）注射到鸡胚发育中的脊髓中，随后进行电穿孔以使dsRNA被摄取。通过这种方法，可以实现基因沉默的时间和空间控制。注射时间点应根据基因的表达谱或蛋白质的半衰期来选择。周转缓慢的蛋白质可能需要在更早阶段进行RNA干扰，理想情况是在基因表达开始之前。可以调整电穿孔参数，以便仅针对脊髓中的特定神经元群体。
