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Plant Cell, Tissue and Organ Culture (PCTOC)

, Volume 120, Issue 2, pp 571–584 | Cite as

The apoplastic oxidative burst as a key factor of hyperhydricity in garlic plantlet in vitro

  • Jie Tian
  • Fangling Jiang
  • Zhen Wu
Original Paper

Abstract

The phenomenon of hyperhydricity, a physiological disorder occurring frequently in tissue culture, causes ultrastructural modification and metabolic alteration of shoots. Reactive oxygen species (ROS) accumulation and oxidative stress induction are common features during the development of hyperhydricity, but the relationship between organelle redox homeostasis and hyperhydricity with ultrastructural abnormalities is unclear. To investigate the origin of oxidative stress-induced hyperhydricity, changes in oxygen metabolism in different subcellular compartments of garlic plantlets in vitro were studied. Under exogenous hydrogen peroxide (H2O2) stress, the chloroplastic and mitochondrial ultrastructure was disrupted, which was concomitant with aggravated frequency and severity of hyperhydricity. The addition of H2O2 to the growth medium enhanced superoxide anion generation and H2O2 content in the subcellular compartments. Accumulation of ROS was the highest in apoplasts. Compared with control shoots, in apoplasts exogenous H2O2 stimulated a sharp increase in superoxide dismutase activity within 4 days and a sharp increase in ascorbate peroxidase and glutathione reductase activities and in ascorbic acid and glutathione contents after 8 days of H2O2 treatment. In the other subcellular compartments, dramatic improvement of the antioxidant system occurred after 12 days. Thus, the apoplast was the most sensitive compartment among those investigated. Apoplastic ROS might play a signaling role to participate in the coordination of stress adaptation. The apoplastic oxidative burst in garlic plantlets in vitro is an early response to the development of hyperhydricity.

Keywords

Garlic Plantlet in vitro Reactive oxygen species Subcellular compartments Hyperhydricity 

Abbreviations

APX

Ascorbate peroxidase

AsA

Ascorbic acid

BSA

Bovine serum albumin

CAT

Catalase

DTNB

5′,5′-Dithiobis-2-nitrobenzoic acid

DTT

dl-Dithiothreitol

EDTA

Ethylene diamine tetraacetic acid

EGTA

Ethylene glycol tetraacetic acid

GR

Glutathione reductase

GSH

Glutathione

GSSG

Oxidized glutathione

HEPES

4-(2-Hydroxyethyl)piperazine-1-ethanesulfonic acid

H2O2

Hydrogen peroxide

IWF

Infiltrated washing fluid

KI

Potassium iodide

NADPH

β-Nicotinamide adenine dinucleotide 2′-phosphate reduced tetrasodium salt

NBT

Nitro blue tetrazolium

O2∙−

Superoxide anion

1O2

Singlet oxygen

OH

Hydroxyl radical

pH

Potential of hydrogen

PMSF

Phenylmethanesulfonyl fluoride

PVP

Polyvinylpyrrolidone

ROS

Reactive oxygen species

SOD

Superoxide dismutase

TCA

Trichloroacetic acid

Notes

Acknowledgments

This work was supported by National Natural Science Foundation of China (31372056) and Doctoral Fund of Ministry of Education of China (200803071012).

Supplementary material

11240_2014_623_MOESM1_ESM.doc (74 kb)
Supplementary material 1 (DOC 74 kb)

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Copyright information

© Springer Science+Business Media Dordrecht 2014

Authors and Affiliations

  1. 1.College of Horticulture, Nanjing Agricultural University/Key Laboratory of Biology and Germplasm Enhancement of Horticultural Crops in East ChinaMinistry of AgricultureNanjingPeople’s Republic of China

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