Biggar Kyle K, Wu Cheng-Wei, Storey Kenneth B
Institute of Biochemistry and Department of Biology, Carleton University, Ottawa, Ontario K1S 5B6, Canada.
Institute of Biochemistry and Department of Biology, Carleton University, Ottawa, Ontario K1S 5B6, Canada.
Anal Biochem. 2014 Oct 1;462:32-4. doi: 10.1016/j.ab.2014.05.032. Epub 2014 Jun 11.
This study makes a significant advancement on a microRNA amplification technique previously used for expression analysis and sequencing in animal models without annotated mature microRNA sequences. As research progresses into the post-genomic era of microRNA prediction and analysis, the need for a rapid and cost-effective method for microRNA amplification is critical to facilitate wide-scale analysis of microRNA expression. To facilitate this requirement, we have reoptimized the design of amplification primers and introduced a polyadenylation step to allow amplification of all mature microRNAs from a single RNA sample. Importantly, this method retains the ability to sequence reverse transcription polymerase chain reaction (RT-PCR) products, validating microRNA-specific amplification.
本研究对先前用于动物模型中无注释成熟微小RNA序列的表达分析和测序的微小RNA扩增技术取得了重大进展。随着研究进入微小RNA预测和分析的后基因组时代，快速且经济高效的微小RNA扩增方法对于促进微小RNA表达的大规模分析至关重要。为满足这一需求，我们重新优化了扩增引物的设计，并引入了聚腺苷酸化步骤，以允许从单个RNA样本中扩增所有成熟微小RNA。重要的是，该方法保留了对逆转录聚合酶链反应（RT-PCR）产物进行测序的能力，从而验证了微小RNA特异性扩增。
