Comparison of Two MicroRNA Quantification Methods for Assaying MicroRNA Expression Profiles in Wheat (Triticum aestivum L.)

doi:10.1016/S2095-3119(13)60362-7

Journal of Integrative Agriculture ›› 2014, Vol. 13 ›› Issue (4): 733-740.DOI: 10.1016/S2095-3119(13)60362-7

Comparison of Two MicroRNA Quantification Methods for Assaying MicroRNA Expression Profiles in Wheat (Triticum aestivum L.)

HAN Ran, YAN Yan, ZHOU Peng , ZHAO Hui-xian

State Key Laboratory of Crop Stress Biology for Arid Areas/College of Life Sciences, Northwest A&F University, Yangling 712100, P.R.China

收稿日期:2013-01-28 出版日期:2014-04-01 发布日期:2014-04-16
通讯作者: ZHAO Hui-xian, Tel: +86-29-87092387, Fax: +86-29-87092262, E-mail: hxzhao212@nwsuaf.edu.cn
作者简介:HAN Ran
基金资助:
This study was financially supported by the National Natural Science Foundation of China (30871578).

Comparison of Two MicroRNA Quantification Methods for Assaying MicroRNA Expression Profiles in Wheat (Triticum aestivum L.)

HAN Ran, YAN Yan, ZHOU Peng , ZHAO Hui-xian

State Key Laboratory of Crop Stress Biology for Arid Areas/College of Life Sciences, Northwest A&F University, Yangling 712100, P.R.China

Received:2013-01-28 Online:2014-04-01 Published:2014-04-16
Contact: ZHAO Hui-xian, Tel: +86-29-87092387, Fax: +86-29-87092262, E-mail: hxzhao212@nwsuaf.edu.cn
About author:HAN Ran
Supported by:
This study was financially supported by the National Natural Science Foundation of China (30871578).

摘要/Abstract

摘要： Two microRNA (miRNA) quantification methods, namely, poly(A) reverse transcription (RT)-quantitative real-time polymerase chain reaction (qPCR) and stem-loop RT-qPCR, have been developed for quantifying miRNA expression. In the present study, five miRNAs, including miR166, miR167, miR168, miR159, and miR396, with different sequence frequencies, were selected as targets to compare their expression profiles in five wheat tissues by applying the two methods and deep sequencing. The study aimed to determine a simple, reliable and high-throughput method for detecting miRNA expressions in wheat tissues. Results showed that the miRNA expression profiles determined by poly(A) RT-qPCR were more consistent with those obtained by deep sequencing. Further analysis indicated that the correlation coefficients of the data obtained by poly(A) RT-qPCR and deep sequencing (0.739, P 0.01) were higher than those obtained by stem-loop RT-qPCR and deep sequencing (0.535, P 0.01). The protocol used for poly(A) RT-qPCR is simpler than that for stem-loop RT-qPCR. Thus, poly(A) RT-qPCR was a more suitable high-throughput assay for detecting miRNA expression profiles. To the best of our knowledge, this study was the first to compare these two miRNA quantification methods. We also provided useful information for quantifying miRNA in wheat or other plant species.

关键词: Triticum aestivum L. , microRNA , deep sequencing , poly(A) RT-qPCR , stem-loop RT-qPCR

Abstract: Two microRNA (miRNA) quantification methods, namely, poly(A) reverse transcription (RT)-quantitative real-time polymerase chain reaction (qPCR) and stem-loop RT-qPCR, have been developed for quantifying miRNA expression. In the present study, five miRNAs, including miR166, miR167, miR168, miR159, and miR396, with different sequence frequencies, were selected as targets to compare their expression profiles in five wheat tissues by applying the two methods and deep sequencing. The study aimed to determine a simple, reliable and high-throughput method for detecting miRNA expressions in wheat tissues. Results showed that the miRNA expression profiles determined by poly(A) RT-qPCR were more consistent with those obtained by deep sequencing. Further analysis indicated that the correlation coefficients of the data obtained by poly(A) RT-qPCR and deep sequencing (0.739, P 0.01) were higher than those obtained by stem-loop RT-qPCR and deep sequencing (0.535, P 0.01). The protocol used for poly(A) RT-qPCR is simpler than that for stem-loop RT-qPCR. Thus, poly(A) RT-qPCR was a more suitable high-throughput assay for detecting miRNA expression profiles. To the best of our knowledge, this study was the first to compare these two miRNA quantification methods. We also provided useful information for quantifying miRNA in wheat or other plant species.

Key words: Triticum aestivum L. , microRNA , deep sequencing , poly(A) RT-qPCR , stem-loop RT-qPCR

HAN Ran, YAN Yan, ZHOU Peng , ZHAO Hui-xian. Comparison of Two MicroRNA Quantification Methods for Assaying MicroRNA Expression Profiles in Wheat (Triticum aestivum L.)[J]. Journal of Integrative Agriculture, 2014, 13(4): 733-740.

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Comparison of Two MicroRNA Quantification Methods for Assaying MicroRNA Expression Profiles in Wheat (Triticum aestivum L.)

Comparison of Two MicroRNA Quantification Methods for Assaying MicroRNA Expression Profiles in Wheat (Triticum aestivum L.)

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