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Overexpression of the Gibberellin 2-Oxidase Gene from Camellia lipoensis Induces Dwarfism and Smaller Flowers in Nicotiana tabacum

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Abstract

Gibberellins (GAs) are plant hormones that control many aspects of plant growth and development. Gibberellin 2-oxidase plays an important role in determining the level of bioactive GAs. In this study, we isolated three GA2ox genes (ClGA2ox1-3) from Camellia lipoensis Chang et Xu. The results of a quantitative real-time reverse transcription polymerase chain reaction analysis indicated that ClGA2ox1-3 may play a tissue-specific role in plant development. The transcript of ClGA2ox1 was more abundant in the stem and apex, ClGA2ox2 was highly expressed in mature leaves, and ClGA2ox3 was more abundant in roots. We produced transgenic plants of Nicotiana tabacum L. by overexpressing the ClGA2ox1-3 genes. Plants with overexpressed ClGA2ox1 or ClGA2ox3 genes exhibited dwarf phenotypes, including reduced growth, delayed flowering, and smaller, rounder, and darker green leaves. All of the transgenic plants overexpressing the ClGA2ox1 gene bloomed normally, but their flowers were half the size of the control plants. Plants overexpressing ClGA2ox3 could be categorized into two classes: moderately dwarfed and severely dwarfed. The ClGA2ox2 gene had little effect on the morphological characterization of transgenic plants. Quantitative real-time PCR analysis showed that the ClGA2ox3 expression level was generally correlated with the level of dwarfism. The endogenous level of bioactive GA4 and GA1 largely decreased in transgenic plants and was generally correlated with the degree of dwarfism in transgenic plants with the ClGA2ox1 or ClGA2ox3 gene. The application of GA3 rescued the dwarf phenotype of transformants, indicating that the GA signaling pathway might function normally in transgenic plants. Therefore, morphological changes in transgenic plants may result from a decrease in the endogenous level of bioactive GAs. Additionally, the possibility of molecular breeding for plant form alternation in Camellia plants by genetically engineering the GA metabolic pathway is discussed.

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Acknowledgments

This work was supported by the National Science & Technology Pillar Program during the 12th Five-year Plan Period (No. 2012BA01B0703) and International Cooperation in Science and Technology Project (No. 2011DFA30490). We also acknowledge Breeding New Flower Varieties Program of Zhejiang Province (2012C12909-6) and State Forestry Administration of the People’s Republic of China, Project 948 (2014-4-16).

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    Authors and Affiliations

    1. Research Institute of Subtropical Forestry/Key Laboratory of Tree Breeding of Zhejiang Province, Chinese Academy of Forestry, Fuyang, Zhejiang, 311400, China

      Zheng Xiao, Jiyuan Li, Zhengqi Fan & Hengfu Yin

    2. Institute of Horticulture, Jiangsu Academy of Agricultural Sciences/Jiangsu Key Laboratory for Horticultural Crop Genetic Improvement, Nanjing, Jiangsu, 210014, China

      Zheng Xiao

    3. College of Enology, Northwest A&F University, Yangling, Shanxi, 712100, China

      Ruipeng Fu

    Authors
    1. Zheng Xiao
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    3. Jiyuan Li
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    Corresponding author

    Correspondence to Jiyuan Li.

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    Zheng Xiao and Ruipeng Fu contributed equally to this work.

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    Figure S1

    Comparison of the deduced amino acid sequences of ClGA2ox1-3 with other GA2oxs. The closed triangles indicate the three amino acid residues forming the iron-binding site. The asterisks show the putative 2-oxoglutarate binding site. The GenBank accession numbers for each protein are ClGA2ox1 (KJ502290), ClGA2ox2 (KJ502289), ClGA2ox3 (KJ502291), AtGA2ox1 (CAB41007), OsGA2ox3 (BAC16752) and PtGA2ox1 (EEE85235). (GIF 182 kb)

    High Resolution Image (TIFF 381 kb)

    Table S1

    Primers used for amplifying GA2ox gene fragment (DOC 30 kb)

    Table S2

    Primers used for RT-PCR analysis (DOC 31 kb)

    Table S3

    Primers used for quantitative real-time PCR (DOC 29 kb)

    Table S4

    Detective primers for southern blot (DOC 28 kb)

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    Xiao, Z., Fu, R., Li, J. et al. Overexpression of the Gibberellin 2-Oxidase Gene from Camellia lipoensis Induces Dwarfism and Smaller Flowers in Nicotiana tabacum . Plant Mol Biol Rep 34, 182–191 (2016). https://doi.org/10.1007/s11105-015-0917-3

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