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Identification of reference genes for tissue-specific gene expression in Panax notoginseng using quantitative real-time PCR

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Abstract

Validated internal controls are prerequisites to accurately normalize gene expression levels. Here, 14 candidate reference genes in Panax notoginseng were characterized. Primer specificity and amplification efficiency were evaluated for each gene. Candidates were subjected to transcript quantification in the root, fibrous root, rhizome, leaf, receptacle, pedicel, and fruit tissues. Expression stability (M value) and normalization factor variation (Vn/Vn+1) were determined by geNorm. 26S-2 and ACT-2 exhibited the highest expression stability among the tissues. Gene expression of dammarenediol synthase was accurately detected after normalization to 26S-2 and ACT-2 was performed. Results were consistent when each or both of 26S-2 and ACT-2 were applied as internal control. Hence, this study provides useful information to normalize gene expression accurately in the tissue-specific transcripts of P. notoginseng.

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Acknowledgments

This work was financially supported by the National Natural Science Foundation of China (NSFC, Grant No. 81260619) and the Scientific Research Project of Guangxi Educational Department (Grant No. 201204LX256). We thank Dr. Hui Yao (Peking Union Medical College, China), Ms. Juan Wang and Ms. Xiaoli Fan (Guilin Medical University, China) for their assistance in the experiments.

Supporting information

Supplementary Table 1—The RT-PCR primer sequences, amplicon size (bp), length of cDNA sequence and accession no. of the candidate genes in P. notoginseng

Supplementary Table 2—The slope, Y-Inter, R2 and amplification efficiency (Eff %) for candidate genes

Supplementary Figure 1—Standard curves of six reference genes and DDS gene were generated from qRT-PCR analyses

Supplementary Figure 2—Average expression stability (M) of six selected reference genes measured by geNorm values

Supplementary Figure 3—Optimal number of reference genes for normalization determined by the pairwise variation V

Author information

Authors and Affiliations

  1. Guangdong Ecological Engineering Vocational College, Guangdong Forestry Vocational Technology School, Guangzhou, 510520, China

    Qiong Wu & Xuehua Feng

  2. Research Center of Chinese Herbal Resource Science and Engineering, Key Laboratory of Chinese Medicinal Resource from Lingnan, Ministry of Education, Guangzhou University of Chinese Medicine, Guangzhou, 510006, China

    Xinye Ma

  3. College of Pharmacy, Guilin Medical University, Guilin, 541004, China

    Kefeng Zhang

Authors
  1. Qiong Wu
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  2. Xinye Ma
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  3. Kefeng Zhang
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  4. Xuehua Feng
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Corresponding author

Correspondence to Qiong Wu.

Electronic supplementary material

Below is the link to the electronic supplementary material.

The online version of this article contains supplementary material, which is available to authorized users. The slope, Y-Inter, R 2 and amplification efficiency (Eff %) for candidate genes, the RT-PCR primer sequences, amplicon size, and accession number of the candidate genes, standard curves of genes and the optimal number of reference genes for normalization are available as Supporting Information.

Supplementary material 1 (DOC 323 kb)

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Wu, Q., Ma, X., Zhang, K. et al. Identification of reference genes for tissue-specific gene expression in Panax notoginseng using quantitative real-time PCR. Biotechnol Lett 37, 197–204 (2015). https://doi.org/10.1007/s10529-014-1643-x

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  • DOI: https://doi.org/10.1007/s10529-014-1643-x

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