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Isolation and characterization of the fragrant cyclamen O-methyltransferase involved in flower coloration

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Abstract

Anthocyanin O-methyltransferase (OMT) is one of the key enzymes for anthocyanin modification and flower pigmentation. We previously bred a novel red-purple-flowered fragrant cyclamen (KMrp) from the purple-flowered fragrant cyclamen ‘Kaori-no-mai’ (KM) by ion-beam irradiation. Since the major anthocyanins in KMrp and KM petals were delphinidin 3,5-diglucoside and malvidin 3,5-diglucoside, respectively, inactivation of a methylation step in the anthocyanin biosynthetic pathway was indicated in KMrp. We isolated and compared OMT genes expressed in KM and KMrp petals. RT-PCR analysis revealed that CkmOMT2 was expressed in the petals of KM but not in KMrp. Three additional CkmOMTs with identical sequences were expressed in petals of both KM and KMrp. Genomic PCR analysis revealed that CkmOMT2 was not amplified from the KMrp genome, indicating that ion-beam irradiation caused a loss of the entire CkmOMT2 region in KMrp. In vitro enzyme assay demonstrated that CkmOMT2 catalyzes the 3′ or 3′,5′ O-methylation of the B-ring of anthocyanin substrates. These results suggest that CkmOMT2 is functional for anthocyanin methylation, and defective expression of CkmOMT2 is responsible for changes in anthocyanin composition and flower coloration in KMrp.

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Abbreviations

OMT:

O-Methyltransferase

Dp3,5dG:

Delphinidin 3,5-diglucoside

Mv3,5dG:

Malvidin 3,5-diglucoside

Pt3,5dG:

Petunidin 3,5-diglucoside

SAM:

S-Adenosyl-l-methionine

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Acknowledgments

We thank N. Sasaki and Y. Ozeki, Tokyo University of Agriculture and Technology, for helpful comments on the biochemical analysis. This work was supported by a grant from the Research and Development Program for New Bio-industry Initiatives of the Bio-oriented Technology Research Advancement Institution (BRAIN).

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Authors and Affiliations

  1. Quantum Beam Science Directorate, Japan Atomic Energy Agency, 1233 Watanuki, Takasaki, Gunma, 370-1292, Japan

    Yusuke Akita, Satoshi Kitamura, Yoshihiro Hase, Issay Narumi & Atsushi Tanaka

  2. Horticultural Laboratory, Saitama Prefecture Agriculture and Forestry Research Center, 91 Rokumanbu, Kuki, Saitama, 346-0037, Japan

    Hiroshi Ishizaka, Emiko Kondo & Naoko Kameari

  3. National Institute of Floricultural Science, National Agriculture and Food Research Organization, 2-1 Fujimoto, Tsukuba, Ibaraki, 305-8519, Japan

    Masayoshi Nakayama, Natsu Tanikawa & Yasumasa Morita

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  1. Yusuke Akita
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  2. Satoshi Kitamura
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  3. Yoshihiro Hase
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  4. Issay Narumi
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  5. Hiroshi Ishizaka
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  8. Masayoshi Nakayama
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  9. Natsu Tanikawa
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  11. Atsushi Tanaka
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Correspondence to Yusuke Akita.

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Akita, Y., Kitamura, S., Hase, Y. et al. Isolation and characterization of the fragrant cyclamen O-methyltransferase involved in flower coloration. Planta 234, 1127–1136 (2011). https://doi.org/10.1007/s00425-011-1466-0

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