Plant Cell, Tissue and Organ Culture (PCTOC)

, Volume 108, Issue 2, pp 237–249 | Cite as

The effects of reactive nitrogen and oxygen species on the regeneration and growth of cucumber cells from isolated protoplasts

  • Marek Petřivalský
  • Pavla Vaníčková
  • Markéta Ryzí
  • Božena Navrátilová
  • Jana Piterková
  • Michaela Sedlářová
  • Lenka Luhová
Original Paper

Abstract

The present study investigated changes in the production of reactive oxygen species (ROS) and reactive nitrogen species (RNS) in isolated mesophyll protoplasts and cell cultures of the cucumber Cucumis sativus cv. Marketer. Although only a minor increase in the level of nitrogen oxide (NO) was observed during the first 7 days of culture following protoplast isolation, a substantial accumulation of ROS was detected. Compounds known to modulate endogenous ROS and RNS levels were employed to study their role in cucumber protoplast regeneration and growth. Supplementing the culture medium with the NO donors S-nitrosoglutathione and sodium nitroprusside and the ROS scavenger ascorbate significantly increased protoplast viability and cell density. In contrast, cell density was significantly decreased following the addition of catalase to the medium. Scavenging of ROS and RNS induced the formation of cucumber microcalli, thus suggesting a differential role of NO in the maintenance of cell viability and in the control of cell division. Our findings confirm the crucial role of controlled ROS and RNS production in both protoplast regeneration and cellular growth and differentiation.

Keywords

Cucumis sativus Protoplast Regeneration Nitric oxide Reactive oxygen species 

Abbreviations

BAP

6-Benzylaminopurine

cPTIO

2-(4-Carboxy-2-phenyl)-4,4,5,5-tetramethylimidazoline-1-oxyl-3-oxide

CW

Calcofluor White

DAF-FM DA

4-Amino-5-methylamino-2′,7′-difluorofluorescein diacetate

FDA

Fluorescein diacetate

GSNO

S-nitrosoglutathione

H2DCF DA

2′,7′-Dichlorofluorescein diacetate

IBA

Indole-3-butyric acid

L-NAME

NG-nitro-l-arginine methyl ester

L-NMMA

NG-monomethyl-l-arginine

MES

2-(N-Morpholino)ethanesulphonic Acid

MTT

3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide

NAA

1-Naphthaleneacetic acid

NO

Nitric oxide

NOS

Nitric oxide synthase

RNS

Reactive nitrogen species

ROS

Reactive oxygen species

SNP

Sodium nitroprusside

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Copyright information

© Springer Science+Business Media B.V. 2011

Authors and Affiliations

  • Marek Petřivalský
    • 1
  • Pavla Vaníčková
    • 1
  • Markéta Ryzí
    • 1
  • Božena Navrátilová
    • 2
  • Jana Piterková
    • 1
  • Michaela Sedlářová
    • 2
  • Lenka Luhová
    • 1
  1. 1.Department of Biochemistry, Faculty of SciencePalacký UniversityOlomoucCzech Republic
  2. 2.Department of Botany, Faculty of SciencePalacký UniversityOlomoucCzech Republic

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