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Transient gene expression in western white pine using agroinfiltration

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Abstract

A method for transient gene expression was developed for western white pine (WWP, Pinus monticola Dougl. ex D.Don) using reporter gene uidA encoding β-glucuronidase (GUS). GUS was transiently expressed in cross sections of primary and secondary needles, cotyledons, and current and second year stems of WWP via vacuum-infiltration with Agrobacterium tumefaciens. Histochemical assays of cross sections of secondary needles showed stronger blue color indicating GUS expression at day 1 and 2 than on other days post agroinfiltration (dpa). GUS activity expressed inside WWP cells was confirmed using light microscopy. In fluorometric assays, GUS expression was high at 1 dpa and lasted until 4 dpa in detached secondary needles, while similarly high expression levels only lasted until 2 dpa in attached secondary needles then dropped significantly. Although the length of GUS-staining zones varied among different WWP organs and between growth and dormant seasons, all tested WWP tissues using the protocol had high levels of transient GUS expression. Thus, heterologous candidate genes or endogenous silencing can be expressed in various WWP tissues or organs using this agroinfiltration approach. The current protocol for efficient transient gene expression will aid functional genomics study of WWP and its pathogens and related conifer species.

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Abbreviations

GUS:

β-glucuronidase

MU:

4-methylumbelliferone

MUG:

4-methylumbelliferyl-β-d-glucuronide

WWP:

Western white pine

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Acknowledgements

Authors appreciate Terry Holmes at CFS for help with microscopy and Gary Roke for providing WWP seedlings.

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Authors and Affiliations

  1. Canadian Forest Service, Natural Resources Canada, 506 West Burnside Road, Victoria, BC, V8Z 1M5, Canada

    Zhenguo Ma, Jun-Jun Liu, Arezoo Zamany & Holly Williams

Authors
  1. Zhenguo Ma
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  2. Jun-Jun Liu
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  3. Arezoo Zamany
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  4. Holly Williams
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Contributions

The project was conceived by JJL. The experiments were designed and performed by ZM. Data were analyzed by ZM with the contribution of JJL, AZ, and HW. The manuscript was written by ZM and JJL with the contribution of AZ and HW.

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Correspondence to Jun-Jun Liu.

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Project funding: The work was partly supported by the CFS-GRDI fund.

The online version is available at http://www.springerlink.com

Corresponding editor: Tao Xu.

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Ma, Z., Liu, JJ., Zamany, A. et al. Transient gene expression in western white pine using agroinfiltration. J. For. Res. 31, 1823–1832 (2020). https://doi.org/10.1007/s11676-019-00938-5

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  • DOI: https://doi.org/10.1007/s11676-019-00938-5

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