基于RNAscope的成年果蝇全脑mRNA多重成像方案。

Protocol for multiplexed RNAscope-based imaging of mRNAs in whole-mount adult Drosophila brains.

作者信息

Wu Meilin, Lambatan Vanessa, Guo Peng, Joiner William J

机构信息

Department of Pharmacology, University of California, San Diego, La Jolla, CA 92093, USA.

Department of Cellular and Molecular Medicine, University of California, San Diego, La Jolla, CA 92093, USA; Nikon Imaging Center, University of California, San Diego, La Jolla, CA 92093, USA.

出版信息

STAR Protoc. 2025 Mar 21;6(1):103615. doi: 10.1016/j.xpro.2025.103615. Epub 2025 Jan 31.

DOI:10.1016/j.xpro.2025.103615

PMID:39891913

原文链接:https://pmc.ncbi.nlm.nih.gov/articles/PMC11835639/

Abstract

Visualizing the expression of mRNAs using traditional in situ hybridization is often hampered by obstacles including weak signal, high background, and poor probe specificity. Here, we present a protocol utilizing RNAscope (ACD) to overcome these obstacles and detect multiple types of mRNAs simultaneously in whole-mount adult Drosophila brains. We further describe how mRNAs can be reliably quantified in any cells that can be targeted by common binary expression systems such as Gal4/UAS and labeled by immunohistochemistry. For complete details on the use and execution of this protocol, please refer to De et al..

摘要

使用传统的原位杂交技术可视化mRNA的表达常常受到诸如信号微弱、背景高和探针特异性差等障碍的阻碍。在这里，我们提出一种利用RNAscope（ACD）的方案来克服这些障碍，并在成年果蝇全脑标本中同时检测多种类型的mRNA。我们进一步描述了如何在任何可被常见二元表达系统（如Gal4/UAS）靶向并通过免疫组织化学标记的细胞中可靠地定量mRNA。有关此方案的使用和执行的完整详细信息，请参考De等人的研究。