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Expression of flavonoid 6-hydroxylase candidate genes in normal and mutant soybean genotypes for glycitein content

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Abstract

Soybean seeds accumulate large amounts of isoflavones (genistein, daidzein and glycitein), secondary metabolites known for their phytoestrogenic activities. Isoflavone composition depends on the seed part and glycitein is almost found exclusively in hypocotyls. Moreover, two major phenotypes are encountered in soybean cultivars, with either low (35 %) or high (55 %) levels of glycitein in their hypocotyls. This trait was under a quasi-mendelian heredity, implicating at most one or two genes. A CYP71D9 cDNA displaying a flavonoid 6-hydroxylase (F6H) activity had previously been isolated from elicitor-induced soybean (Glycine max L.) cells. This enzyme allows the synthesis of the glycitein flavanone intermediate (6,7,4′- trihydroxyflavanone) by catalyzing the A-ring hydroxylation of liquiritigenin. In this study, the CYP71D9 gene (F6H1) and two other candidates (F6H2 and F6H3) were studied using contrasted soybean cultivars for glycitein content (0, 35, 55 and 80 %). Their expression was observed in chitosan elicited leaves. They encode P450 proteins of 496, 469 and 481 amino acids respectively and were expressed in leaves with or without elicitation. The expression patterns of these three genes were performed in cotyledons and hypocotyls at different developmental stages. F6H1 and F6H2 were not expressed in the developing seed. F6H3 was only expressed in hypocotyls. Its expression levels did not correlate with hypocotyls glycitein content, but it was not expressed in the null mutant for glycitein. Thus, this F6H3 gene is a good potential candidate for glycitein biosynthesis in soybean seed.

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Acknowledgments

We thank Christel Couderc and Daphné Puech for their help and technical assistance. This project was supported by research grants from the Midi-Pyrénées Région. The authors thank Pr Randall Nelson, curator of the USDA soybean germplasm collection (Urbana, IL), for the supply of the accession used in this study and the French group Euralis Semences for the supply of their germplasm accessions NS Kasha.

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  1. Toxalim UMR 1331 INRA/INP/UPS, Ecole d’Ingénieurs Purpan, Université de Toulouse, 75, voie du TOEC, BP 57611, 31076, Toulouse Cedex 3, France

    Marie-Pierre Artigot, Mathieu Baes, Jean Daydé & Monique Berger

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  1. Marie-Pierre Artigot
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  2. Mathieu Baes
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  3. Jean Daydé
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Correspondence to Monique Berger.

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Artigot, MP., Baes, M., Daydé, J. et al. Expression of flavonoid 6-hydroxylase candidate genes in normal and mutant soybean genotypes for glycitein content. Mol Biol Rep 40, 4361–4369 (2013). https://doi.org/10.1007/s11033-013-2526-2

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  • DOI: https://doi.org/10.1007/s11033-013-2526-2

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