Effect of UV-C irradiation on mesophyll protoplasts of Cucumis sativus
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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.
KeywordsAsymmetric fusion Calcofluor White Cell wall regeneration Cucumis sativus DAPI FDA Mesophyll protoplast
Murashige and Skoog
α-Naphthalene acetic acid
Relative fluorescence unit
This work was supported by the Grant MSM 6198959215 from the Ministry of Education, Youth and Sports of the Czech Republic and by the project NAZV QF 4108. We would like to thank to Research Institute of Crop Production (Prague, Czech Republic) and to Department of Gene Bank (Olomouc, Czech Republic) for providing of seed material.
- Atanassov A, Dimantov D, Atanassov I, Dragoeva A, Vassileva Z, Vitanov V, Jankulova M, Djilianov D (1991) Transfer of resistance to tomato spotted wilt virus from wild Nicotiana species to N. tabacum via somatic hybridization. Physiol Plant 82:A23, poster session 8, No. 130Google Scholar
- Gajdová J, Lebeda A, Navrátilová B (2004) Protoplast cultures of Cucumis and Cucurbita spp. In: Lebeda A, Paris HS (eds) Progress in cucurbit genetics and breeding research. Proceedings of Cucurbitaceae 2004, 8th EUCARPIA meeting on cucurbit genetics and breeding, Palacký University in Olomouc, Olomouc, Czech Republic, 2004, pp 441–454Google Scholar
- Gajdová J, Navrátilová B, Smolná J, Lebeda A (2007) Factors affecting protoplast isolation and cultivation of Cucumis spp. J Appl Bot Food Qual 81:1–6Google Scholar
- Seemüller E (1976) Investigation to demonstrate mycoplasma-like organism in diseased plants by fluorescence microscopy. Acta Hortic 67:109–112Google Scholar