In Vitro Cellular & Developmental Biology - Plant

, Volume 54, Issue 6, pp 642–657 | Cite as

Validation of reference genes for gene expression analysis of response to anthocyanin induction in cell cultures of Vitis davidii (Rom. Caill.) Foëx

  • Chengchun LaiEmail author
  • Hong Pan
  • Xiangui Huang
  • Lihua Fan
  • Changqing Duan
  • Shaozhen Li
Plant Tissue Culture


Real-time quantitative polymerase chain reaction (RT-qPCR) is an effective method for detecting changes of gene expression in plant cell metabolic regulation. A set of 15 reference gene candidates were selected for the present study of anthocyanin biosynthesis regulation, and stability. The suitability of their expression was evaluated in eight different experimental treatments in spine grape (Vitis davidii [Rom. Caill.] Foëx.) cell cultures. The results indicated that SAND family protein (SAND) and V-type proton ATPase subunit G (VAG) were the most stable reference genes for culture duration, tubulin alpha-3/alpha-5 chain (α-tubulin) and tubulin beta-1 chain (β-tubulin) for illumination conditions, ubiquitin-conjugating enzyme E2-17 kDa (UBQ) and VAG for UVB treatment, VAG and 60S ribosomal protein L18-2 (60SRP) for temperature treatment, AP47, clathrin adaptor complex subunit mu (AP-2) and 60SRP for cinnamic acid treatment, α-tubulin and UBQ for chitosan treatment, actin and alcohol dehydrogenase 2 (ADH2) for kinetin treatment, and β-tubulin and elongation factor 1-α (EF1-α) for cell line. Finally, the reliability of the selected reference genes was confirmed by investigating the expression profiles of the target gene dihydroflavonol 4-reductase (DFR) in spine grape cell cultures. The results of the present study offer the most robust platform for the most precise and broad application of RT-qPCR to investigate gene expression associated with anthocyanin biosynthesis in spine grape cell cultures.


Spine grape (Vitis davidii [Rom. Caill.] Foëx.) Cell culture Gene expression Reference gene Real-time quantitative PCR 


Funding information

This work was supported by Fujian provincial natural science fund subject (no. 2016J01126), Fujian Provincial Department of Science and Technology of special public-funded projects (no. 2016R1014-1), and FAAS Scientific and Technological Innovation Team (grant no. STIT2017-1-10).


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Copyright information

© The Society for In Vitro Biology 2018

Authors and Affiliations

  • Chengchun Lai
    • 1
    • 2
    • 3
    Email author
  • Hong Pan
    • 1
  • Xiangui Huang
    • 1
  • Lihua Fan
    • 1
  • Changqing Duan
    • 2
  • Shaozhen Li
    • 3
  1. 1.Institute of Agricultural Engineering and TechnologyFujian Academy of Agricultural SciencesFuzhouChina
  2. 2.Center for Viticulture and Enology, College of Food Science & Nutritional EngineeringChina Agricultural UniversityBeijingChina
  3. 3.Beijing Huiyuan Food & Beverage Co., Ltd.BeijingChina

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