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Hairy Root-activation Tagging: a High-throughput System for Activation Tagging in Transformed Hairy Roots

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Abstract

Activation tagging is a powerful technique for generating gain-of-function mutants in plants. We developed a new vector system for activation tagging of genes in “transformed hairy roots”. The binary vector pHR-AT (Hairy Root-Activation Tagging) and its derivative pHR-AT-GFP contain a cluster of rol (rooting locus) genes together with the right border facing four tandem repeats of the cauliflower mosaic virus (CaMV) 35S enhancer element on the same T-DNA. Transformation experiments using Arabidopsis, potato, and tobacco as model plants revealed that upon inoculating plants with Agrobacterium tumefaciens harboring these vectors, a large number of independently transformed roots could be induced from explants within a short period of time, and root culture lines were subsequently established. Molecular analyses of the pHR-AT-GFP-transformed Arabidopsis lines showed that expression of the genes adjacent to the T-DNA insertion site was significantly increased. This system may facilitate application of the activation-tagging approach to plant species that are recalcitrant to the regeneration of transgenic plants. High-throughput metabolic profiling of activation-tagged root culture lines will offer opportunities for identifying regulatory or biosynthetic genes for the production of valuable secondary metabolites of interest.

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Authors and Affiliations

  1. RIKEN Plant Science Center, Kanagawa, Tsurumi-ku, Yokohama, 230-0045, Japan

    Hikaru Seki, Tomoko Nishizawa, Shigeo Yoshida & Toshiya Muranaka

  2. Center for Gene Science , Hiroshima University, Kagamiyama, Higashi-Hiroshima, Hiroshima, 739-8527, Japan

    Nobukazu Tanaka

  3. Laboratory of Plant Cell Technology, Graduate School of Nutritional and Environmental Sciences, University of Shizuoka, 52-1 Yada, Shizuoka, 422-8526, Japan

    Yasuo Niwa

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  1. Hikaru Seki
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  2. Tomoko Nishizawa
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  3. Nobukazu Tanaka
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  4. Yasuo Niwa
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  5. Shigeo Yoshida
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  6. Toshiya Muranaka
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Correspondence to Toshiya Muranaka.

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Seki, H., Nishizawa, T., Tanaka, N. et al. Hairy Root-activation Tagging: a High-throughput System for Activation Tagging in Transformed Hairy Roots. Plant Mol Biol 59, 793–807 (2005). https://doi.org/10.1007/s11103-005-1008-x

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