Plant Cell, Tissue and Organ Culture

, Volume 94, Issue 3, pp 313–318 | Cite as

Effect of UV-C irradiation on mesophyll protoplasts of Cucumis sativus

  • Božena Navrátilová
  • Lenka Luhová
  • Marek Petřivalský
Original Paper

Abstract

In the present work we studied the effect of UV-C irradiation on short-term protoplast physiology, with the aim of identifying and assessing parameters which can provide valuable information for asymmetric fusion experiments. Protoplast viability, cell wall regeneration, density of cell suspension and intensity of DAPI signal were followed by using microscopy and by the detection of specific fluorescent or spectroscopic signals in a microplate reader. The control and irradiated mesophyll protoplasts of Cucumis sativus were used for this experiment. In contrast to control cells, viability of irradiated cells significantly decreased. Intensive cell wall regeneration was observed only in control cells, which also showed significantly higher DAPI fluorescence signal. Microscopy for determination of viability by FDA and cell wall regeneration by Calcofluor White were modified for microplate reader instrumentation. These methods are simple, fast and suitable for detection of the effectiveness of UV-C irradiation of cells intended to be used in asymmetric fusion experiments.

Keywords

Asymmetric fusion Calcofluor White Cell wall regeneration Cucumis sativus DAPI FDA Mesophyll protoplast 

Abbreviations

BAP

Benzylaminopurine

CW

Calcofluor White

DAPI

4′,6-Diamidino-2-phenylindole

FDA

Fluorescein diacetate

IBA

Indole-3-butyric acid

LM

Liquid medium

MES

2-(N-morpholino)ethanesulfonic acid

MS

Murashige and Skoog

NAA

α-Naphthalene acetic acid

PS

Protoplast suspension

RFU

Relative fluorescence unit

UV

Ultraviolet light

2,4-D

2,4-Dichlorophenoxyacetic acid

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Copyright information

© Springer Science+Business Media B.V. 2008

Authors and Affiliations

  • Božena Navrátilová
    • 1
  • Lenka Luhová
    • 2
  • Marek Petřivalský
    • 2
  1. 1.Department of Botany, Faculty of SciencePalacký University in OlomoucOlomouc-HoliceCzech Republic
  2. 2.Department of Biochemistry, Faculty of SciencePalacký University in OlomoucOlomouc-HoliceCzech Republic

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