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3 Biotech

, 8:148 | Cite as

A simplified and efficient Agrobacterium tumefaciens electroporation method

  • Evelin Kámán-Tóth
  • Miklós Pogány
  • Tamás Dankó
  • Ágnes Szatmári
  • Zoltán Bozsó
Short Reports

Abstract

Agrobacterium tumefaciens is a widely used microbial tool in plant molecular biology to transfer DNA into plant cells and produce, e.g., stable or transient transformants or induce gene silencing. In our study, we present a simplified version of electrocompetent cell preparation that is not only time and cost efficient, but it requires minimal handling of bacterial cells. Liquid cultures are normally used to prepare competent Agrobacterium cells. To overcome the difficulties of working with liquid cultures, we propose suspending bacterial cells directly from overnight agar plate cultures. In addition, we optimized several parameters to simplify the procedure and maximize the number of transformants (e.g., Agrobacterium strains, number of washing steps, amount of required plasmid DNA, electroporation parameters, type of incubation media, or incubation time). This optimized, simple, and fast protocol has proved to be efficient enough to obtain transformed colonies with low amounts (as little as 1 ng) of plasmid DNA. In addition, it also enabled us to introduce ligated plasmids directly into Agrobacterium omitting the E. coli transformation step and accelerating the cloning procedure further.

Keywords

Agrobacterium Electroporation Plate method Direct transformation 

Notes

Acknowledgements

Research in the authors’ laboratories has been funded by the Hungarian Scientific Research Fund (OTKA K 104730, OTKA PD 109050), the Bolyai Scholarship (BO 609 12, BO 00348/09), and the Széchenyi 2020 Programme, the European Regional Development Fund and the Hungarian Government (GINOP-2.3.2-15-2016-00061) which are gratefully acknowledged.

Compliance with ethical standards

Conflict of interest

The authors declare that they have no conflicts of interest in the publication.

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Copyright information

© Springer-Verlag GmbH Germany, part of Springer Nature 2018

Authors and Affiliations

  1. 1.Department of Pathophysiology, Centre for Agricultural Research, Plant Protection InstituteHungarian Academy of SciencesBudapestHungary

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