Center for Life Nano Science@Sapienza, Istituto Italiano di Tecnologia, Rome, Italy.
Department of Biology and Biotechnology Charles Darwin, Sapienza University of Rome, Rome, Italy.
Methods Mol Biol. 2021;2157:251-280. doi: 10.1007/978-1-0716-0664-3_15.
The RNA fluorescence in situ hybridization (RNA-FISH) methodology offers an attractive strategy to deepen our knowledge on the long noncoding RNA biology. In this chapter, we provide a comprehensive overview of the current RNA-FISH protocols available for imaging nuclear and cytoplasmic lncRNAs within cells or tissues. We describe a multicolor approach optimized for the simultaneous visualization of these transcripts with their specific molecular interactors, such as proteins or DNA sequences. Common challenges faced by this methodology such as cell-type specific permeabilization, target accessibility, image acquisition, and post-acquisition analyses are also discussed.
RNA 荧光原位杂交（RNA-FISH）方法为深入了解长非编码 RNA 生物学提供了一种有吸引力的策略。在本章中，我们提供了一个全面的概述，介绍了目前可用于在细胞或组织内成像核和细胞质 lncRNA 的 RNA-FISH 方案。我们描述了一种多色方法，该方法经过优化，可同时可视化这些转录本与其特定的分子相互作用物（如蛋白质或 DNA 序列）。还讨论了该方法面临的常见挑战，如细胞类型特异性通透、靶标可及性、图像采集和采集后分析。
