Owens Geoffrey C, Edelman David B
Department of Neurosurgery, David Geffen School of Medicine at the University of California, Los Angeles, 300 Stein Plaza, Ste. 562, Los Angeles, CA, 90095, USA,
Methods Mol Biol. 2015;1313:237-46. doi: 10.1007/978-1-4939-2703-6_18.
Mitochondria are highly dynamic organelles that undergo fusion and fission on a relatively fast time scale. Here, a straightforward method is described for capturing mitochondrial fusion events in real time using a photoconvertible fluorescent protein and a far-field fluorescence microscope equipped with appropriate image acquisition and analysis software. The Kaede photoconvertible fluorescent protein is tagged with a mitochondrial targeting sequence and delivered to primary neurons by lentiviral transduction, which ensures efficient low copy number transgene insertion, as well as stable transgene expression.
线粒体是高度动态的细胞器，在相对较快的时间尺度上发生融合和裂变。本文描述了一种直接的方法，可使用光转换荧光蛋白和配备适当图像采集与分析软件的远场荧光显微镜实时捕获线粒体融合事件。Kaede光转换荧光蛋白用线粒体靶向序列进行标记，并通过慢病毒转导递送至原代神经元，这确保了低拷贝数转基因的高效插入以及转基因的稳定表达。
