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Reference gene selection for qRT-PCR in Caragana korshinskii Kom. under different stress conditions

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Abstract

Caragana korshinskii Kom., which is widely distributed in the northwest China and Mongolia, is an important forage bush belonging to the legume family with high economic and ecological value. Strong tolerance ability to various stresses makes C. korshinskii Kom. a valuable species for plant stress research. In this study, suitable reference genes for quantitative real-time reverse transcription PCR (qRT-PCR) were screened from 11 candidate reference genes, including ACT, GAPDH, EF1α, UBQ, TUA, CAP, TUB, TUB3, SKIP1, SKIP5-1 and SKIP5-2. A total of 129 samples under drought, heat, cold, salt, ABA and high pH treatment were profiled, and software such as geNORM, NormFinder and BestKeeper were used for reference gene evaluation and selection. Different suitable reference genes were selected under different stresses. Across all 129 samples, GAPDH, EF1α and SKIP5-1 were found to be the most stable reference genes, and EF1α+SKIP5-1 is the most stable reference gene combination. Conversely, TUA, TUB and SKIP1 were not suitable for using as reference genes owing to their great expression variation under some stress conditions. The relative expression levels of CkWRKY1 were detected using the stable and unstable reference genes and their applicability was confirmed. These results provide some stable reference genes and reference gene combinations for qRT-PCR under different stresses in C. korshinskii Kom. for future research work, and indicate  that CkWRKY1 plays essential roles in response to stresses in C. korshinskii.

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Abbreviations

qRT-PCR:

Reverse transcription quantitative real-time PCR

ACT:

Actin

GAPDH:

Glyceraldehyde-3-phosphate dehydrogenase

EF1α:

Elongation factor 1-alpha

UBQ:

Ubiquitin

TUA:

Alpha-tubulin

CAP:

Adenylyl cyclase-associated protein

TUB:

Beta-tubulin

TUB3:

Tubulin beta-3

SKIP1:

SKP1/ASK-interacting protein 1

SKIP5-1/SKIP5-2:

SKP1/ASK-interacting protein 5

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Acknowledgments

We deeply appreciate Dr. Mark Goettel, Editor-in-Chief of Biocontrol Science & Technology, for polishing the manuscript carefully. This work was financially supported by National Natural Science Foundation of China (No. 31360169), Chinese National Programs for High Technology Research and Development (No. 2011AA100203), the Program for New Century Excellent Talents of University (No. ECNT-11-1020) from Ministry of Education (China), and the Innovative Research Group Fund (No. NDPYTD2010-3) from Inner Mongolia Agricultural University (China).

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The authors declare that they have no conflict of interest.

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Authors and Affiliations

  1. College of Life Sciences, Inner Mongolia Agricultural University, Hohhot, 010018, People’s Republic of China

    Qi Yang, Jiajia Yin, Gao Li, Ruigang Wang & Guojing Li

  2. Research Institute of Forestry, Chinese Academy of Forestry, Beijing, 100091, People’s Republic of China

    Liwang Qi

  3. College of Food Science and Bioengineering, Inner Mongolia Agricultural University, Hohhot, 010018, People’s Republic of China

    Feiyun Yang

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  1. Qi Yang
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Correspondence to Guojing Li.

Additional information

Qi Yang, Jiajia Yin and Gao Li contributed equally to this work.

Key message: The stable reference genes for gene expression evaluation of Caragana korshinskii under drought, heat, cold, salt, ABA and high pH treatments were selected respectively from 11 candidate genes.

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Yang, Q., Yin, J., Li, G. et al. Reference gene selection for qRT-PCR in Caragana korshinskii Kom. under different stress conditions. Mol Biol Rep 41, 2325–2334 (2014). https://doi.org/10.1007/s11033-014-3086-9

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