Comprehensive evaluation of extracellular small RNA isolation methods from serum in high throughput sequencing.
Comprehensive evaluation of extracellular small RNA isolation methods from serum in high throughput sequencing.
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DOI：
10.1186/s12864-016-3470-z
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发表时间：
2017-01-07
期刊：
影响因子：
4.4
通讯作者：
Flynn CR
中科院分区：
文献类型：
--
作者：
Guo Y;Vickers K;Xiong Y;Zhao S;Sheng Q;Zhang P;Zhou W;Flynn CR
来自全血、血清和血浆的DNA和RNA是越来越受欢迎的分析物，目前正在研究它们在疾病诊断和分期中的用途。小的非编码核糖核酸（srna），特别是microrna （mirna）及其变体异构体（isomir），以及转移RNA （tRNA）衍生的小RNA （tdr）构成了在这方面特别有希望的分子库。在这项设计的研究中，我们比较了分离循环细胞外sRNAs（前sRNAs）的各种方法和试剂盒的性能。使用五种不同的分离试剂盒：Qiagen循环核酸试剂盒、ThermoFisher Scientific Ambion TRIzol LS试剂、Qiagen miRNEasy、QiaSymphony RNA提取试剂盒和Exiqon MiRCURY RNA分离试剂盒，从一个健康个体中分离前srnas。每种分离方法重复4次。共构建了20个小RNA测序（sRNAseq）文库，并采用严格的生物信息学方法进行了测序和比较。循环核酸试剂盒具有最大的miRNA分离变异性，但对其他RNA类别（isomiRs， tdr和其他杂项sRNAs (osRNA)）的分离变异性最低。然而，与表现最好的方法Ambion TRIzol相比，循环核酸试剂盒始终产生最少的基因组测序，后者绘制了10%的mirna， 7.2%的tdr和23.1%的osRNAs。其他方法发挥了中间作用，QiaSymphony绘制了14%的osRNAs， miRNEasy绘制了4.6%的tdr和2.9%的mirna，总体上获得了第二好的试剂盒性能评级。总之，根据下游应用和需要处理的样品数量，每个分离试剂盒显示出不同的性能特征，这些特征可能被解释为偏向或有利。本文的在线版本（doi:10.1186/s12864-016-3470-z）包含补充材料，授权用户可以使用。
DNA and RNA fractions from whole blood, serum and plasma are increasingly popular analytes that are currently under investigation for their utility in the diagnosis and staging of disease. Small non-coding ribonucleic acids (sRNAs), specifically microRNAs (miRNAs) and their variant isoforms (isomiRs), and transfer RNA (tRNA)-derived small RNAs (tDRs) comprise a repertoire of molecules particularly promising in this regard. In this designed study, we compared the performance of various methods and kits for isolating circulating extracellular sRNAs (ex-sRNAs). ex-sRNAs from one healthy individual were isolated using five different isolation kits: Qiagen Circulating Nucleic Acid Kit, ThermoFisher Scientific Ambion TRIzol LS Reagent, Qiagen miRNEasy, QiaSymphony RNA extraction kit and the Exiqon MiRCURY RNA Isolation Kit. Each isolation method was repeated four times. A total of 20 small RNA sequencing (sRNAseq) libraries were constructed, sequenced and compared using a rigorous bioinformatics approach. The Circulating Nucleic Acid Kit had the greatest miRNA isolation variability, but had the lowest isolation variability for other RNA classes (isomiRs, tDRs, and other miscellaneous sRNAs (osRNA). However, the Circulating Nucleic Acid Kit consistently generated the fewest number of reads mapped to the genome, as compared to the best-performing method, Ambion TRIzol, which mapped 10% of the miRNAs, 7.2% of the tDRs and 23.1% of the osRNAs. The other methods performed intermediary, with QiaSymphony mapping 14% of the osRNAs, and miRNEasy mapping 4.6% of the tDRs and 2.9% of the miRNAs, achieving the second best kit performance rating overall. In summary, each isolation kit displayed different performance characteristics that could be construed as biased or advantageous, depending upon the downstream application and number of samples that require processing. The online version of this article (doi:10.1186/s12864-016-3470-z) contains supplementary material, which is available to authorized users.
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影响因子：
14.9
作者：
Flicek P;Amode MR;Barrell D;Beal K;Billis K;Brent S;Carvalho-Silva D;Clapham P;Coates G;Fitzgerald S;Gil L;Girón CG;Gordon L;Hourlier T;Hunt S;Johnson N;Juettemann T;Kähäri AK;Keenan S;Kulesha E;Martin FJ;Maurel T;McLaren WM;Murphy DN;Nag R;Overduin B;Pignatelli M;Pritchard B;Pritchard E;Riat HS;Ruffier M;Sheppard D;Taylor K;Thormann A;Trevanion SJ;Vullo A;Wilder SP;Wilson M;Zadissa A;Aken BL;Birney E;Cunningham F;Harrow J;Herrero J;Hubbard TJ;Kinsella R;Muffato M;Parker A;Spudich G;Yates A;Zerbino DR;Searle SM
通讯作者：
Searle SM
影响因子：
14.9
作者：
Buschmann D;Haberberger A;Kirchner B;Spornraft M;Riedmaier I;Schelling G;Pfaffl MW
通讯作者：
Pfaffl MW
影响因子：
7
作者：
Berezikov, Eugene;Robine, Nicolas;Lai, Eric C.
通讯作者：
Lai, Eric C.
血清 microRNA 是有前途的弥漫性大 B 细胞淋巴瘤的新型生物标志物
影响因子：
3.5
作者：
Fang, Cheng;Zhu, Dan-Xia;Li, Jian-Yong
通讯作者：
Li, Jian-Yong
影响因子：
6.3
作者：
Guo Y;Xiong Y;Sheng Q;Zhao S;Wattacheril J;Flynn CR
通讯作者：
Flynn CR