A non-radioactive method for small RNA detection by northern blotting.

A non-radioactive method for small RNA detection by northern blotting.

Small non-coding RNAs are essential regulators of gene expression at the transcriptional and posttranscriptional levels. High-throughput sequencing has revealed thousands of predicted small RNAs; however, only a few of these have been well characterized. Northern blotting is the most convincing method for small RNA validation. In this study, we improved the Northern blot method by using biotin-labeled probes. miRNAs and siRNAs derived from both Arabidopsis thaliana and Oryza sativa were investigated. The results suggest that this improved method is sensitive and efficient, with approximately 5 μg of total RNA being sufficient for detection. Furthermore, long-term storage of probes labeled in this manner is more convenient, less contaminative and degradative compared with traditional probes. This protocol is an alternative strategy for small RNA detection and represents an efficient means of researching small RNAs. The online version of this article (doi:10.1186/s12284-014-0026-1) contains supplementary material, which is available to authorized users.

小非编码RNA是转录水平和转录后水平基因表达的重要调节因子。高通量测序已经揭示了数千种预测的小RNA；然而，其中只有少数得到了很好的表征。Northern杂交是小RNA验证最可靠的方法。 在这项研究中，我们通过使用生物素标记的探针改进了Northern杂交方法。对来自拟南芥和水稻的微小RNA（miRNA）和小干扰RNA（siRNA）进行了研究。结果表明，这种改进的方法灵敏且高效，大约5μg总RNA就足以进行检测。此外，与传统探针相比，以这种方式标记的探针长期储存更方便，污染和降解更少。 该实验方案是小RNA检测的一种替代策略，是研究小RNA的一种有效手段。 本文的网络版（doi:10.1186/s12284 - 014 - 0026 - 1）包含补充材料，授权用户可获取。