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Optimization of Agrobacterium-mediated transient expression of heterologous genes in spinach

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Abstract

The Agrobacterium-mediated transient assay is a relatively rapid technique and a promising approach for assessing the expression of a gene of interest. Despite the successful application of this transient expression system in several plant species, it is not well understood in spinach. In this study, we analyzed various factors, including infiltration method, Agrobacterium strain and density, and co-infiltration of an RNA silencing suppressor (p19), that affect transient expression following agroinfiltration in spinach. To evaluate the effects of these factors on the transient expression system, we used the β-glucuronidase (GUS) reporter gene construct pB7WG2D as a positive control. The vacuum-based infiltration method was much more effective at GUS gene expression than was the syringe-based infiltration method. Among the three Agrobacterium strains examined (EHA105, LBA4404, and GV2260), infiltration with the GV2260 strain suspension at a final optical cell density (OD600) of 1.0 resulted in the highest gene expression. Furthermore, co-expression of suppressor p19 also increased the efficiency and duration of gene expression and protein accumulation. The results indicate that the use of optimized conditions for transient gene expression could be a simple, rapid, and effective tool for functional genomics in spinach.

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Acknowledgements

This work was conducted with the support of the “Cooperative Research Program for Agriculture Science & Technology Development (Project No. PJ011289032017)”, the Rural Development Administration, the Republic of Korea and the Basic Science Research Program through the National Research Foundation of Korea (NRF), funded by the Ministry of Education (2016R1D1A1B02012307 and 2016R1A6A1A03012862).

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  1. Dang Viet Cao and Reniel S. Pamplona contributed equally to this research.

Authors and Affiliations

  1. Department of Biotechnology, College of Applied Life Science (SARI), Jeju National University, Jeju, 63243, South Korea

    Dang Viet Cao, Reniel S. Pamplona, Young Kyoung Oh, Key-Zung Riu & Kyung-Hwan Boo

  2. Woojung Institute of Life Sciences, Woojung BSC, Seongnam, Gyeonggi, 16229, South Korea

    Jiwon Kim & Jongcheol Ahn

  3. Department of Systems Biology, College of Life Science and Biotechnology, Yonsei University, Seoul, 03722, South Korea

    Seok-Keun Cho & Seong-Wook Yang

  4. Department of Plant and Environmental Sciences, Faculty of Science, University of Copenhagen, Thorvaldsensvej 40, 1871, Copenhagen, Denmark

    Seong-Wook Yang

  5. Subtropical/tropical Organism Gene Bank, Jeju National University, Jeju, 63243, South Korea

    Key-Zung Riu & Kyung-Hwan Boo

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  1. Dang Viet Cao
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Correspondence to Key-Zung Riu or Kyung-Hwan Boo.

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Cao, D.V., Pamplona, R.S., Kim, J. et al. Optimization of Agrobacterium-mediated transient expression of heterologous genes in spinach. Plant Biotechnol Rep 11, 397–405 (2017). https://doi.org/10.1007/s11816-017-0457-4

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