Direct S-Poly(T) Plus assay in quantification of microRNAs without RNA extraction and its implications in colorectal cancer biomarker studies
Direct S-Poly(T) Plus assay in quantification of microRNAs without RNA extraction and its implications in colorectal cancer biomarker studies
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无需提取 RNA 即可直接进行 S-Poly(T) Plus 定量 microRNA 测定及其在结直肠癌生物标志物研究中的意义
DOI：
10.1186/s12967-019-2061-6
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发表时间：
2019-09
期刊：
影响因子：
--
通讯作者：
Deming Gou
中科院分区：
文献类型：
--
作者：
Yanqin Niu;Sijian Xia;Mingyang Su;Quanjin Dang;Kang Kang;Li Li;Deming Gou
研究背景微小RNAs(MiRNAs)生物标记物的研究进展产生了具有潜在临床价值的疾病标记物。然而，直到今天，这些已发表的结果都没有应用于临床。主要原因可能是缺乏简单而可靠的miRNA检测方法。方法我们建立了一种简单但超灵敏的RT-qPCR方法，即直接S-Poly(T)Plus法，该方法无需纯化RNA即可检测miRNA。本研究对该方法进行了优化，并与其他基于RNA纯化的miRNA检测方法进行了比较，并对其灵敏度进行了测试。利用直接S-Poly(T)Plus方法对7个潜在的结直肠癌miRNA生物标志物进行了验证。结果该方法可以在最少的血浆输入(20μL)和最短的时间(~140min)下检测到大约100个miRNA。该方法的灵敏度是茎环法的2.7~343倍，与S-保利(T)Plus法相当。用直接S-Poly(T)PLUS方法验证的7个结直肠癌miRNA生物标志物可以区分I期结直肠癌和健康人，与受试者工作特征(ROC)曲线下面积在0.79~0.94(p值0.001)之间具有良好的区分性。结论该方法简便、可靠，可用于临床特异性miRNAs的研究。
BackgroundAdvances in microRNAs (miRNAs) biomarkers have generated disease markers with potential clinical values. However, none of these published results have been applied in clinic until today. The main reason could be the lack of simple but robust miRNA measurements.MethodsWe built up a simple but ultrasensitive RT-qPCR protocol, Direct S-Poly(T) Plus assay, for detecting miRNAs without RNA purification. In this study, the method was optimized and compared with other RNA purification-based miRNA assays, and the sensitivity was tested. Using Direct S-Poly(T) Plus method, seven potential miRNA biomarkers of colorectal cancer were validated.ResultsIt is possible to detect approximately 100 miRNAs with minimal plasma inputs (20 μl) and time (~ 140 min) with this approach. The sensitivity of this method was 2.7–343-fold higher than that of the stem-loop method, and comparable with S-Poly(T) plus method. 7 validated miRNA biomarkers of colorectal cancer by Direct S-Poly(T) plus assay could discriminate colorectal cancer stage I from healthy individuals, and promised satisfactory discrimination with the area under receiver operating characteristic (ROC) curve ranging from 0.79 to 0.94 (pvalue 0.001).ConclusionsThis simple and robust protocol may have strong impact on the development of specific miRNAs as biomarkers in clinic.
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影响因子：
4.7
作者：
Resnick, Kimberly E.;Alder, Hansjuerg;Cohn, David E.
通讯作者：
Cohn, David E.
影响因子：
14.9
作者：
Chen C;Ridzon DA;Broomer AJ;Zhou Z;Lee DH;Nguyen JT;Barbisin M;Xu NL;Mahuvakar VR;Andersen MR;Lao KQ;Livak KJ;Guegler KJ
通讯作者：
Guegler KJ
DOI：
10.1186/1758-907x-1-7
发表时间：
2010-03-01
期刊：
Silence
影响因子：
--
作者：
Kosaka N;Izumi H;Sekine K;Ochiya T
通讯作者：
Ochiya T
影响因子：
3.7
作者：
Ho YK;Xu WT;Too HP
通讯作者：
Too HP
影响因子：
56.9
作者：
Lim, LP;Glasner, ME;Bartel, DP
通讯作者：
Bartel, DP