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Cloning and characterization of a functional flavanone-3ß-hydroxylase gene from Medicago truncatula

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Abstract

As a key enzyme in the biosynthesis of flavonols, anthocyanidins and proanthocyanidins, flavanone-3ß-hydroxylase (F3H) plays very important roles in plant stress response. A putative flavanone-3ß-hydroxylase gene from Medicago truncatula (MtF3H), a model legume species, was identified from a bio-data analysis platform. It was speculated to be induced by salt stress based on the outcomes of the analysis platform. The complementary DNA (cDNA) consists of 1499 bp with an open reading frame (ORF) of 1098 bp, which encodes a putative protein of 365 amino acids with a molecular weight of about 41.36 kDa and an isoelectric point of 5.60. To measure the catalytic activity of the protein, the MtF3H gene was ligated to pYES2 vector and heterologously expressed in yeast. The recombinant protein converted naringen into dihydrokaempferol and displayed different enzymatic efficiencies with other flavanones, confirming that MtF3H coding a functional flavanone-3ß-hydroxylase. The expression pattern of the MtF3H gene was analyzed by comparative quantitative RT-PCR and a higher level of expression was observed in the roots than was observed in stems and leaves. Furthermore, the expression was induced by salt stress in the roots, and to a greater extent in the stems, but the response of the gene activity to salt stress in the stems was slower in the first 12 h following treatment when compared to the roots.

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Acknowledgments

This work was supported by the Hi-Tech Research and Development (863) Program of China (2006AA10Z105, 2006AA100109). We thank Dr. Dasharath Lohar for providing seeds of M. truncatula Gaertn ‘Jemalong’ A17. We also sincerely thank Dr. shuizhang Fei (Iowa state University, Ames, USA) for his help.

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Authors and Affiliations

  1. State Key Laboratory for Agro-biotechnology, College of Biological Sciences, China Agricultural University, Beijing, 100193, People’s Republic of China

    Xiaoye Shen, Mingliang Chen, Daofeng Li, Jiangli Dong & Tao Wang

  2. Philipps-Universität Marburg, Institut für Pharmazeutische Biologie, Deutschhausstr. 17A, 35037, Marburg/Lahn, Germany

    Stefan Martens

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  1. Xiaoye Shen
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Correspondence to Jiangli Dong.

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Shen, X., Martens, S., Chen, M. et al. Cloning and characterization of a functional flavanone-3ß-hydroxylase gene from Medicago truncatula . Mol Biol Rep 37, 3283–3289 (2010). https://doi.org/10.1007/s11033-009-9913-8

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