核糖核酸
活体细胞成像
生物
细胞生物学
绿色荧光蛋白
荧光显微镜
基因表达
转染
基因
细胞
荧光
遗传学
物理
量子力学
作者
Jae Youn Shim,Byung Hun Lee,Hye Yoon Park
出处
期刊:Methods in molecular biology
日期:2019-01-01
卷期号:: 47-61
被引量:2
标识
DOI:10.1007/978-1-4939-9674-2_4
摘要
Transcription and post-transcriptional regulations are critical in gene expression. To study the spatiotemporal regulation of RNA inside a cell, techniques for high-resolution imaging of RNA have been developed. In this chapter, we describe RNA fluorescent labeling methods using MS2 and PP7 systems to detect single RNA molecules in live neurons. We use hippocampal neurons cultured from knock-in mouse models in which β-actin or Arc mRNAs are tagged with MS2 or PP7 stem-loops. Adeno-associated virus (AAV) or lentiviral vectors are used to express MS2 or PP7 capsid proteins fused with GFP in those neurons. Then, GFP-labeled RNAs in live neurons can be detected by epifluorescence microscopy, and their moving pathways can be analyzed using single-particle tracking software. For these processes, we introduce protocols for neuron culture, transfection, imaging, and particle tracking methods.
科研通智能强力驱动
Strongly Powered by AbleSci AI