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CLC number: S823.9+1

On-line Access: 2015-06-08

Received: 2014-12-14

Revision Accepted: 2015-04-29

Crosschecked: 2015-05-20

Cited: 0

Clicked: 1969

Citations:  Bibtex RefMan EndNote GB/T7714

 ORCID:

Hong-yun Liu

http://orcid.org/0000-0001-8806-6204

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Journal of Zhejiang University SCIENCE B 2015 Vol.16 No.6 P.533-540

http://doi.org/10.1631/jzus.B1400355


Comparative studies of two methods for miRNA isolation from milk whey


Author(s):  Xiao-lu Jin, Zi-hai Wei, Lan Liu, Hong-yun Liu, Jian-xin Liu

Affiliation(s):  Institute of Dairy Science, MoE Key Laboratory of Molecular Animal Nutrition, College of Animal Sciences, Zhejiang University, Hangzhou 310058, China

Corresponding email(s):   hyliu@zju.edu.cn, liujx@zju.edu.cn

Key Words:  Method, Milk whey miRNA, Spike-in miRNA


Xiao-lu Jin, Zi-hai Wei, Lan Liu, Hong-yun Liu, Jian-xin Liu. Comparative studies of two methods for miRNA isolation from milk whey[J]. Journal of Zhejiang University Science B, 2015, 16(6): 533-540.

@article{title="Comparative studies of two methods for miRNA isolation from milk whey",
author="Xiao-lu Jin, Zi-hai Wei, Lan Liu, Hong-yun Liu, Jian-xin Liu",
journal="Journal of Zhejiang University Science B",
volume="16",
number="6",
pages="533-540",
year="2015",
publisher="Zhejiang University Press & Springer",
doi="10.1631/jzus.B1400355"
}

%0 Journal Article
%T Comparative studies of two methods for miRNA isolation from milk whey
%A Xiao-lu Jin
%A Zi-hai Wei
%A Lan Liu
%A Hong-yun Liu
%A Jian-xin Liu
%J Journal of Zhejiang University SCIENCE B
%V 16
%N 6
%P 533-540
%@ 1673-1581
%D 2015
%I Zhejiang University Press & Springer
%DOI 10.1631/jzus.B1400355

TY - JOUR
T1 - Comparative studies of two methods for miRNA isolation from milk whey
A1 - Xiao-lu Jin
A1 - Zi-hai Wei
A1 - Lan Liu
A1 - Hong-yun Liu
A1 - Jian-xin Liu
J0 - Journal of Zhejiang University Science B
VL - 16
IS - 6
SP - 533
EP - 540
%@ 1673-1581
Y1 - 2015
PB - Zhejiang University Press & Springer
ER -
DOI - 10.1631/jzus.B1400355


Abstract: 
MicroRNAs (miRNAs) from milk whey have been considered for their potential as noninvasive biomarkers for milk quality control and disease diagnosis. However, standard protocols for miRNA isolation and quantification from milk whey are not well established. The objective of this study was to compare two methods for the isolation of miRNAs from milk whey. These two methods were modified phenol-based technique (Trizol LS® followed by phenol precipitation, the TP method) and combined phenol and column-based approach (Trizol LS® followed by cleanup using the miRNeasy kit, the TM method). Yield and quality of RNA were rigorously measured using a NanoDrop ND-1000 spectrophotometer and then the distribution of RNA was precisely detected in a Bioanalyzer 2100 instrument by microchip gel electrophoresis. Several endogenous miRNAs (bta-miR-141, bta-miR-146a, bta-miR-148a, bta-miR-200c, bta-miR-362, and bta-miR-375) and an exogenous spike-in synthetic control miRNA (cel-miR-39) were quantified by real-time polymerase chain reaction (PCR) to examine the apparent recovery efficiency of milk whey miRNAs. Both methods could successfully isolate sufficient small RNA (<200 nt) from milk whey, and their yields were quite similar. However, the quantification results show that the total miRNA recovery efficiency by the TM method is superior to that by the TP method. The TM method performed better than the TP for recovery of milk whey miRNA due to its consistency and good repeatability in endogenous and spike-in miRNA recovery. Additionally, quantitative recovery analysis of a spike-in miRNA may be more accurate to reflect the milk whey miRNA recovery efficiency than using traditional RNA quality analysis instruments (NanoDrop or Bioanalyzer 2100).

两种乳清miRNA分离提取方法的比较研究

目的:系统比较两种从液态乳清中分离提取miRNA的方法。
创新点:首次比较发现提取柱吸附法分离乳清miRNA的效果优于醇沉淀法。
方法:采用柱吸附法和醇沉淀法分别对乳清中的miRNA进行提取,采用NanoDropND-1000分光光度计和基于Bioanalyzer2100的微芯片凝胶电泳检测RNA提取质量。通过荧光定量聚合酶链式反应(PCR)检测外源加入的miRNA及乳清内源的miRNA丰度。确定最优方法后扩大样本容量,采用20份牛奶样本进行提取,并通过荧光定量PCR检测了外源miRNA提取的稳定性及牛奶内源miRNA提取的重复性。
结论:乳清miRNA提取柱吸附法优于醇沉淀法,且重复性好。

关键词:乳清miRNA;提取方法;柱吸附法

Darkslateblue:Affiliate; Royal Blue:Author; Turquoise:Article

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