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Int. J. Mol. Sci. 2015, 16(9), 20468-20491; doi:10.3390/ijms160920468

Evaluation of Appropriate Reference Genes for Reverse Transcription-Quantitative PCR Studies in Different Tissues of a Desert Poplar via Comparision of Different Algorithms

1
National Engineering Laboratory for Tree Breeding, College of Biological Sciences and Technology, Beijing Forestry University, Beijing 100083, China
2
The Key Laboratory for Silviculture and Conservation of Ministry Education, College of Forestry, Beijing Forestry University, Beijing 100083, China
3
Institute of Crop Sciences, Chinese Academy of Agricultural Sciences, Beijing 100081, China
These authors contributed equally to this work.
*
Authors to whom correspondence should be addressed.
Academic Editor: Ann Cuypers
Received: 23 April 2015 / Accepted: 7 August 2015 / Published: 28 August 2015
(This article belongs to the Special Issue Abiotic Stress and Gene Networks in Plants)
View Full-Text   |   Download PDF [1226 KB, uploaded 28 August 2015]   |  

Abstract

Despite the unshakable status of reverse transcription-quantitative PCR in gene expression analysis, it has certain disadvantages, including that the results are highly dependent on the reference genes selected for data normalization. Since inappropriate endogenous control genes will lead to inaccurate target gene expression profiles, the validation of suitable internal reference genes is essential. Given the increasing interest in functional genes and genomics of Populus euphratica, a desert poplar showing extraordinary adaptation to salt stress, we evaluated the expression stability of ten candidate reference genes in P. euphratica roots, stems, and leaves under salt stress conditions. We used five algorithms, namely, ΔCt, NormFinder, geNorm, GrayNorm, and a rank aggregation method (RankAggreg) to identify suitable normalizers. To support the suitability of the identified reference genes and to compare the relative merits of these different algorithms, we analyzed and compared the relative expression levels of nine P. euphratica functional genes in different tissues. Our results indicate that a combination of multiple reference genes recommended by GrayNorm algorithm (e.g., a combination of Actin, EF1α, GAPDH, RP, UBQ in root) should be used instead of a single reference gene. These results are valuable for research of gene identification in different P. euphratica tissues. View Full-Text
Keywords: reference genes; salt stress; normalization; reverse transcription-quantitative PCR; Populus euphratica reference genes; salt stress; normalization; reverse transcription-quantitative PCR; Populus euphratica
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This is an open access article distributed under the Creative Commons Attribution License which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. (CC BY 4.0).

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Wang, H.-L.; Li, L.; Tang, S.; Yuan, C.; Tian, Q.; Su, Y.; Li, H.-G.; Zhao, L.; Yin, W.; Zhao, R.; Xia, X. Evaluation of Appropriate Reference Genes for Reverse Transcription-Quantitative PCR Studies in Different Tissues of a Desert Poplar via Comparision of Different Algorithms. Int. J. Mol. Sci. 2015, 16, 20468-20491.

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