新方法定量血浆外泌体miRNA 或可作为转移性乳腺癌标记物

土豆

文章原文附件见2楼。

已有研究证实，循环miRNA是由核酸酶降解RNA形成的并会进入微膜泡和外泌体从而被组装。量化从这些膜泡中完全释放的miRNA是研究循环miRNA的关键一步。目前基于纯化RNA的定量分析被广泛使用，而且通过这种总RNA的提取和小RNA富集过程得到的这些低丰度的miRNA，是非常消耗时间和成本的。

同济大学医学院的研究人员优化了一种简单、有效和节省时间的方法来直接定量血浆miRNA而不需要进行总RNA的分离纯化。该方法基于miRNA会从蛋白质复合物所完全释放，随后进行反转录和实时定量PCR扩增（RT-PCR）来检测。与传统的直接提取RNA的分析方法相比，这种直接定量方法在循环miRNA分析上，表现出更高的效率、更高的准确性以及特异性。通过这种直接定量方法对临床样品与基于RNA的miRNA的芯片分析相结合的应用中，研究人员验证了血液中的miR-106a在转移性乳腺癌患者会上调，暗示miR-106a可能会是转移性乳腺癌的潜在生物标志物。

图：血浆miRNA的直接定量方法示意图。

参考文献：Zhao Q, Deng S, Wang G, Liu C, Meng L, Qiao S, Shen L, Zhang Y, Lü J, Li W, Zhang Y, Wang M, Pestell RG, Liang C, Yu Z. (2016) A direct quantification method for measuring plasma MicroRNAs identified potential biomarkers for detecting metastatic breast cancer. Oncotarget [Epub ahead of print].

原文摘要：Circulating miRNAs are protected from ribonuclease degradation by assembly into microvesicles and exosomes. Releasing miRNAs completely from these particles is the key step to quantify the circulating miRNAs. Currently purified RNA-based quantitative analysis is widely used while it is time and cost consuming with high risk for those circulating miRNAs with low abundance due to partial loss of RNA during the steps of total RNA extraction and small RNA enrichment. Herein, we optimized a simple, effective and time-saving method to directly measure plasma miRNAs without RNA isolation. It is based on complete miRNA release from the protein complexes, followed by miRNA-specific reverse transcription and quantitative real-time PCR amplification.By comparison to the RNA-based approach, the direct quantification method showed more efficiency for circulating miRNA analysis, higher accuracy and specificity. By
application of the direct quantification method to clinical samples combined with the RNA-based miRNA screening analysis, upregulation of miR-106a in blood was validated in metastatic breast cancer patients, indicating miR-106a are a potential biomarker for metastatic breast cancer.

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土豆 发表于 2016-3-17 17:09
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