一种分析微小RNA（miRNA）表达的简单方法。

A simple method for profiling miRNA expression.

作者信息

Wang Jia-Wang, Cheng Jin Q

机构信息

H. Lee Moffitt Cancer Center and Research Institute, Tampa, FL, USA.

出版信息

Methods Mol Biol. 2008;414:183-90. doi: 10.1007/978-1-59745-339-4_14.

DOI:10.1007/978-1-59745-339-4_14

PMID:18175820

Abstract

Here we describe a simple protocol that uses positively charged nylon membrane dot blot to profile miRNA expression. A library of 515 antisense oligodeoxynucleotides of human and mouse mature miRNAs was synthesized and spotted on GeneScreen Plus membrane using a dot-blot equipment. Total RNA or enriched small molecular weight RNAs (smwRNAs) were enzymatically radiolabeled by poly (A) polymerase and then hybridized to the nylon membrane oligo arrays. The spot signal intensity on the membrane was analyzed using phosphorimaging. This method offers a convenient and economic way to simultaneously detect the expression of hundreds of miRNAs.

摘要

在此，我们描述了一种简单的实验方案，该方案使用带正电荷的尼龙膜斑点印迹法来分析miRNA的表达。合成了一个包含515个人类和小鼠成熟miRNA反义寡脱氧核苷酸的文库，并使用斑点印迹设备将其点样在GeneScreen Plus膜上。总RNA或富集的小分子重量RNA（smwRNAs）通过聚（A）聚合酶进行酶促放射性标记，然后与尼龙膜寡核苷酸阵列杂交。使用磷光成像分析膜上的斑点信号强度。该方法提供了一种方便且经济的方式来同时检测数百种miRNA的表达。