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Generation of stable transgenic Brassica napus cv. Jet Neuf cell cultures as a tool to investigate in planta protein function

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Abstract

Analysis of proteins often requires generation of transgenic plants to gain knowledge of their in planta function. The acquisition of such plants represents a significant investment in terms of workload, time, and space. Here we describe an easy-to-use alternative system by transforming Brassica napus cv. Jet Neuf suspension cells derived from microspores. The cells can be transformed with high efficiency, and even double transformations with different transgenes are possible. In addition, we describe a procedure that allows for long-term storage to maintain diverse transgenic cell lineages with low effort, to build up cell suspension libraries for research purposes. Overall, the work can be helpful for scientists that want to use B. napus as a research organism in a reduced time-frame without the need for generating whole plants and the requirement of larger greenhouse spaces.

Key message

This study establishes a protocol for generation of stable Brassica napus transgenic cell cultures, their long-term storage, and contains examples how these cell cultures can be used in studying gene function in planta without generating whole plants.

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Data availability

The datasets of the current study are presented in this article or are available upon a reasonable request.

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Acknowledgements

We would like to thank Prof. Thomas Clemente, Center for Plant Science Innovation, Department of Agronomy and Horticulture, University of Nebraska-Lincoln, for providing the RD29a promoter. We also would like to thank Dr. John Shanklin, Brookhaven National Laboratory, NY, for providing the stock of B. napus cv. Jet Neuf cell cultures used in this work, and WSU’s Franceschi Microscopy and Imaging Center for providing access to a confocal microscope and supporting undergraduate research.

Funding

This project was supported by an Agriculture and Food Research Initiative competitive grant 2019-67013-29160 of the USDA National Institute of Food and Agriculture (NIFA) to H.H. WSU financially supported J.L. in his undergraduate research studies through a CAS Summer Fellowship.

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Contributions

J.L. is an undergraduate student of WSU’s Honors College who established the transformation and long storage procedures, and also performed the GUS stainings shown in this work. S.M. did the RT-qPCR analysis, S.S. did salt and low-sucrose with salt assays, M.K. did the confocal analysis, and R.A-S. generated the proESR1:GUS and proRD29a:BrRAP2.4-1 expression constructs. H.H. is the principal investigator, designed the figures, and developed the manuscript together with J.L. All authors contributed with editing and agreed on the manuscript.

Corresponding author

Correspondence to Hanjo Hellmann.

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The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.

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Communicated by Heidi Halbwirth.

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Landers, J., Mooney, S., Smalley, S. et al. Generation of stable transgenic Brassica napus cv. Jet Neuf cell cultures as a tool to investigate in planta protein function. Plant Cell Tiss Organ Cult 154, 633–643 (2023). https://doi.org/10.1007/s11240-023-02538-y

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  • DOI: https://doi.org/10.1007/s11240-023-02538-y

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