Affinity Purification of Binding miRNAs for Messenger RNA Fused with a Common Tag

• Main Authors: Ke Wei, Feng Yan, Hui Xiao, Xiaoxu Yang, Guie Xie, Ye Xiao, Tingting Wang, Yu Xun, Zhaoqin Huang, Mei Han, Jian Zhang, Shuanglin Xiang
• Format: Article
• Language: English
• Published: MDPI AG 2014-08-01
• Series: International Journal of Molecular Sciences
• Subjects: affinity purification; mRNA; target miRNA
• Online Access: http://www.mdpi.com/1422-0067/15/8/14753

Prediction of microRNA–mRNA interaction typically relies on bioinformatic methods, but these methods only suggest the possibility of microRNA binding and may miss important interactions as well as falsely predict others. A major obstacle to the miRNA research has been the lack of experimental procedures for the identification of miRNA–mRNA interactions. Recently, a few studies have attempted to explore experimental methods to isolate and identify miRNA targets or miRNAs targeting a single gene. Here, we developed an more convenient experimental approach for the isolation and identification of miRNAs targeting a single gene by applying short biotinylated DNA anti-sense oligonucleotides mix to enhanced green fluorescent protein (EGFP) mRNA which was fused to target gene mRNA. This method does not require a design of different anti-sense oligonucleotides to any mRNA. This is a simple and an efficient method to potentially identify miRNAs targeting specific gene mRNA combined with chip screen.