Simple and Nonradioactive Detection of MicroRNAs Using Digoxigenin (DIG)-labeled Probes with High Sensitivity

Overview

Journal RNA

Specialty Molecular Biology

Date 2014 Feb 28

PMID 24572812

Citations 4

Authors

Wei Wu

Pengtao Gong

Jianhua Li

Ju Yang

Guocai Zhang

He Li

Zhengtao Yang

Xichen Zhang

Affiliations

Soon will be listed here.

Abstract

The discovery of microRNAs (miRNAs), which are ∼21-23 nucleotides that can regulate targeted mRNA by transcript cleavage or protein translation suppression, has changed the landscape of biomedical field greatly. At present, Northern blot analysis based on radioisotopes is still the most popular method on the detection of miRNAs for its high sensitivity. However, radioisotopes have been known for certain disadvantages, such as instability, expense, and safety; thus, developing a nonradioactive and highly sensitive method is needed. Here, we report a simple, nonradioactive, and sensitive method for miRNAs detection based on 5'-phos-3'-DIG-labeled probes prepared through splinted ligation and EDC cross-linking (DSLE). The method was more sensitive than traditional Northern blots with a DIG-labeled DNA probe and can detect as low as 2 fmol of miRNAs. The whole procedure can be completed within 6-8 h. DSLE method is very convenient, cost-effective, time-saving, and highly sensitive.

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