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Virus-induced multiple gene silencing to study redundant metabolic pathways in plants: Silencing the starch degradation pathway in Nicotiana benthamiana

Authors

  • Dr. Gavin M. George,

    Corresponding author
    1. Genetics Department, Institute for Plant Biotechnology, Stellenbosch University, Matieland, South Africa
    • Genetics Department, Institute for Plant Biotechnology, Stellenbosch University, Private Bag X1, Matieland 7602, South Africa
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  • Rolene Bauer,

    1. Genetics Department, Institute for Plant Biotechnology, Stellenbosch University, Matieland, South Africa
    2. Department of Biotechnology, Institute for Microbial Biotechnology and Metagenomics, University of the Western Cape, Bellville, Cape Town, South Africa
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  • Andreas Blennow,

    1. Research Centre Pro-Active Plants, Faculty of Science, Department of Plant Biology and Biotechnology, University of Copenhagen, Denmark
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  • Jens Kossmann,

    1. Genetics Department, Institute for Plant Biotechnology, Stellenbosch University, Matieland, South Africa
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  • James R. Lloyd

    1. Genetics Department, Institute for Plant Biotechnology, Stellenbosch University, Matieland, South Africa
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Abstract

Virus-induced gene silencing (VIGS) is a rapid technique that allows for specific and reproducible post-transcriptional degradation of targeted mRNA. The method has been proven efficient for suppression of expression of many single enzymes. The metabolic networks of plants, however, often contain isoenzymes and gene families that are able to compensate for a mutation and mask the development of a silencing phenotype. Here, we show the application of multiple gene VIGS repression for the study of these redundant biological pathways. Several genes in the starch degradation pathway [disproportionating enzyme 1; (DPE1), disproportionating enzyme 2 (DPE2), and GWD] were silenced. The functionally distinct DPE enzymes are present in alternate routes for sugar export to the cytoplasm and result in an increase in starch production when silenced individually. Simultaneous silencing of DPE1 and DPE2 in Nicotiana benthamiana resulted in a near complete suppression in starch and accumulation of malto-oligosaccharides.

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