Quantitative Analysis of Alternative Pre-mRNA Splicing in Mouse Brain Sections Using RNA <em>In Situ</em> Hybridization Assay

Xuan Guo; Yiqing Zhao; Hieu Nguyen; Tonghua Liu; Zhenghe Wang; Hua Lou

doi:10.3791/57889

JoVE Journal
Genetics

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JoVE Journal Genetics

Quantitative Analysis of Alternative Pre-mRNA Splicing in Mouse Brain Sections Using RNA In Situ Hybridization Assay

RNA原位杂交法定量分析小鼠脑切片中的替代前 mRNA 拼接方法

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11:22 min

August 26, 2018

DOI: 10.3791/57889-v

Xuan Guo^1,2, Yiqing Zhao^1,3, Hieu Nguyen¹, Tonghua Liu², Zhenghe Wang^1,3, Hua Lou^1,3,4

¹Department of Genetics and Genome Sciences,Case Western Reserve University, ²Department of Endocrinology,Dongfang Hospital of Beijing University of Chinese Medicine, ³Case Comprehensive Cancer Center,Case Western Reserve University, ⁴Center for RNA Science and Therapeutics,Case Western Reserve University

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本文介绍了一种利用短反义寡核苷酸检测小鼠脑部替代前 mRNA 拼接模式的原位杂交协议。

这种方法可以帮助回答选择性 pre-mRNA 剪接领域的关键问题。具体来说，它提供了一个稳健、灵敏的检测系统，用于原位检查组织切片中的选择性剪接模式。使用这种方法，可以同时分析复杂生物结构（如小鼠大脑）中包含的许多亚型的选择性剪接。

该技术的主要优点是它在 mRNA 原位杂交中使用休克抗静电外显子连接探针来产生亚型特异性杂交信号。构建信号放大技术提高了检测的灵敏度，以提供稳定的杂交信号。检测到的信号是逃逸特定探针的外显子包涵体，这些探针用于计算小鼠大脑不同解剖区域生命体征剪接的百分比。

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遗传学问题 138 原位杂交神经纤维瘤类型 1 外显子 23a 小鼠脑替代剪接