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Coloring genetically modified soybean grains with anthocyanins by suppression of the proanthocyanidin genes ANR1 and ANR2

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Abstract

Detection and quantification of the levels of adventitious presence of genetically modified (GM) soybeans in non-GM grain shipments currently requires sophisticated tests that can have issues with their reproducibility. We show here that pigment biosynthesis in the soybean seed coat can be manipulated to provide a distinct color that would enable the simple visible detection of the GM soybean grain. We observed that a distinct red-brown grain color could be engineered by the simultaneous suppression of two proanthocyanidin (PA) genes, ANTHOCYANIDIN REDUCTASE1 (ANR1) and ANR2. Multiple reaction monitoring by liquid chromatography tandem mass spectrometry was used to quantify differentially accumulated seed coat metabolites, and revealed the redirection of metabolic flux into the anthocyanin pigment pathway and unexpectedly the flavonol-3-O-glucoside pathway. The upregulations of anthocyanin isogenes (DFR1 and GST26) and the anthocyanin/flavonol-3-O-glycosyltransferase (UGT78K2) were identified by quantitative RT-PCR to be endogenous feedback and feedforward responses to overaccumulation of upstream flavonoid intermediates resulting from ANR1 and ANR2 suppressions. These results suggested the transcription of flavonoid genes to be a key component of the mechanism responsible for the redirection of metabolite flux. This report identifies the suppression of PA genes to be a novel approach for engineering pigmentation in soybean grains.

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Acknowledgments

We gratefully thank Dr. Vincenzo De Luca for the fruitful conversations, Dr. Ko Shimamoto for providing the pANDA35HK vector and Carla Schmidt for assistance with soybean transformation. This project was funded by the A-base (project inventory 126) from Agriculture and Agri-Food Canada to BM and DB, and NSERC Discovery Grants to BM and JTA.

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Authors and Affiliations

  1. Bioproducts and Bioprocesses, Research Branch, Agriculture and Agri-Food Canada, Ottawa, ON, Canada

    Nik Kovinich, Tara L. Rintoul & Brian Miki

  2. Department of Biology, Ottawa–Carleton Institute of Biology, Carleton University, Ottawa, ON, Canada

    Nik Kovinich

  3. Department of Biology and Center for Research in Biopharmaceuticals and Biotechnology, University of Ottawa, Ottawa, ON, Canada

    Ammar Saleem & John T. Arnason

  4. Southern Crop Protection and Food Research Centre, Agriculture and Agri-Food Canada, London, ON, N5V 4T3, Canada

    Daniel C. W. Brown

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  1. Nik Kovinich
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Correspondence to Nik Kovinich.

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Kovinich, N., Saleem, A., Rintoul, T.L. et al. Coloring genetically modified soybean grains with anthocyanins by suppression of the proanthocyanidin genes ANR1 and ANR2 . Transgenic Res 21, 757–771 (2012). https://doi.org/10.1007/s11248-011-9566-y

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  • DOI: https://doi.org/10.1007/s11248-011-9566-y

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