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Practical guidelines for the characterization of tobacco BY-2 cell lines

  • Original Papers
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Biologia Plantarum

Abstract

Plant cell lines represent useful models in plant cell biology. They allow simple analysis of the effects of various factors including modulated gene expression at cellular and subcellular levels. The tobacco BY-2 cell line is a favoured model due to its high proliferation rate, capability of effective synchronization, and accessibility to transformation. A relatively high uniformity of BY-2 cultures allows morphological phenotyping and assessment of growth parameters like mitotic index, viability, or cell density. Here we review already published and newly introduced optimized guidelines to carry out reliable, reproducible and efficient characterization of BY-2 cultures from suggestions of appropriate methods to acquire primary data, proper statistical treatment, and biological interpretation. The presented experimental data demonstrate the extent of natural variability and the effect of initial cell density on various cell culture features. Supportive equations allow to estimate some derived phenotypic parameters like cell cycle duration or fresh biomass of the culture and to determine the size of data sets for reliable documentation of a certain phenotypic change. The optimized protocols and accompanying discussion of weak points of different approaches should serve as practical guide for both beginners and experienced researchers working on BY-2 cells.

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Abbreviations

2,4-D:

2,4-dichlorophenoxyacetic acid

CV:

coefficient of variation

DIC:

differential interference contrast microscopy

FDA:

fluorescein diacetate

MI:

mitotic index

SBI:

subculture interval

SD:

standard deviation

VIA:

viability

WT:

wild type

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Authors and Affiliations

  1. Department of Experimental Plant Biology, Faculty of Science, Charles University in Prague, Viničná 5, CZ-12844, Prague, Czech Republic

    M. Srba, Z. Opatrný & L. Fischer

  2. Institute of Applied Mathematics and Information Technologies, Faculty of Science, Charles University in Prague, Albertov 6, CZ-12843, Prague, Czech Republic

    A. Černíková

Authors
  1. M. Srba
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  2. A. Černíková
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  3. Z. Opatrný
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  4. L. Fischer
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Correspondence to M. Srba.

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Acknowledgement: Funding was provided by the Ministry of Education, Youth and Sports of the Czech Republic (NPUI L01417). We thank Dr. Tomas Urfus for flow-cytometry analysis of BY-2 cells. We also thank Dr. Maria Barker-Mott for linguistic revision of the manuscript.

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Srba, M., Černíková, A., Opatrný, Z. et al. Practical guidelines for the characterization of tobacco BY-2 cell lines. Biol Plant 60, 13–24 (2016). https://doi.org/10.1007/s10535-015-0573-3

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  • DOI: https://doi.org/10.1007/s10535-015-0573-3

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