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Molecular characterization and expression analysis of dihydroflavonol 4-reductase (DFR) gene in Saussurea medusa

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Abstract

Dihydroflavonol 4-reductase (DFR), which catalyzes the reduction of dihydroflavonols to leucoanthocyanins, is a key enzyme in the biosynthesis of anthocyanidins, proanthocyanidins, and other flavonoids of importance in plant development and human nutrition. This study isolated a full length cDNA encoding DFR, designated as SmDFR (GenBank Accession No. EF600682), by screening a cDNA library from a red callus line of Saussurea medusa, which is an endangered, traditional Chinese medicinal plant with high pharmacological value. SmDFR was functionally expressed in yeast (Saccharomyces cerevisiae) to confirm that SmDFR can readily reduce dihydroquercetin (DHQ) and dihydrokampferol (DHK), but it could not reduce dihydromyricetin (DHM). The deduced SmDFR structure shared extensive sequence similarity with previously characterized plant DFRs and phylogenetic analysis showed that it belonged to the plant DFR super-family. SmDFR also possessed flavanone 4-reductase (FNR) activity and can catalyze the conversion of eridictyol to luteoforol. Real-time PCR analysis showed that the expression level of SmDFR was higher in flowers compared with both leaves and roots. This work greatly enhances our knowledge of flavonoid biosynthesis in S. medusa and marks a major advance that could facilitate future genetic modification of S. medusa.

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Abbreviations

ANS:

Anthocyanidin synthase

BA:

Benzyladenine

CHI:

Chalcone isomerase

DHK:

Dihydrokaempferol

DHM:

Dihydromyricetin

DHQ:

Dihydroquercetin

DFR:

Dihydroflavonol 4-reductase

HPLC-MS:

High performance liquid chromatography-mass spectrometry

F3H:

Flavanone 3-hydroxylase

F3′H:

Flavanonoid 3′ hydroxylase

FNR:

Flavanone 4-reductase

NAA:

Naphthaleneacetic Acid

NADPH:

Reduced nicotinamide adenine dinucleotide phosphate

ORF:

Open reading frame

PAL:

Phenylalanine ammonia-lyase

UTR:

Untranslated region

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Acknowledgments

This work was supported by the National Science Foundation of China (No. 30873452; 30701089). The authors thank Dr. Duncan E. Jackson for critical review and comments to improve the manuscript.

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Authors and Affiliations

  1. Key Laboratory of Photosynthesis and Environmental Molecular Physiology, Institute of Botany, Chinese Academy of Sciences, Beijing, 100093, China

    Houhua Li, Jian Qiu, Fudong Chen, Xiaofen Lv, Dexiu Zhao & Xuejun Hua

  2. Forage Improvement Division, The Samuel Roberts Noble Foundation, Ardmore, OK, 73401, USA

    Chunxiang Fu

  3. Plant Biology Division, The Samuel Roberts Noble Foundation, Ardmore, OK, 73401, USA

    Qiao Zhao

  4. College of Forestry, Northwest A&F University, Yangling, 712100, Shaanxi, China

    Houhua Li

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  1. Houhua Li
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  2. Jian Qiu
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Correspondence to Xuejun Hua or Qiao Zhao.

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Li, H., Qiu, J., Chen, F. et al. Molecular characterization and expression analysis of dihydroflavonol 4-reductase (DFR) gene in Saussurea medusa . Mol Biol Rep 39, 2991–2999 (2012). https://doi.org/10.1007/s11033-011-1061-2

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