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Isolation of mitochondria from embryogenic cultures of Picea abies (L.) Karst. and Abies cephalonica Loud.: characterization of a K +ATP channel

  • Physiology and Biochemistry
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Abstract

A valuable method to isolate and purify mitochondria from embryonal masses of two coniferous species (Picea abies [L.] Karst. and Abies cephalonica Loud.) is described. Crude mitochondria from both species were shown to be intact, oxygen consuming (with malate plus glutammate, succinate and NADH as substrates) and well coupled (respiratory control ratio ca. 4). The oxidation of the substrates was only partially KCN-insensitive (alternative oxidase) in some cases. However, these fractions were contaminated by membranes (e.g. plasmalemma, tonoplast, Golgi and endoplasmic reticulum). After purification by a discontinuous Percoll gradient (18, 23, 40%, v/v), three mitochondrial populations were separated. The 0/18 interface fraction was composed mainly of broken and uncoupled mitochondria, while the other two (18/23 and 23/40 interface fractions) contained intact and coupled mitochondria, but only 23/40 interface fraction revealed to be better purified starting from both coniferous embryonal masses. In the latter purified fraction, the presence of a cyclosporin A-sensitive K +ATP channel was demonstrated. These findings were discussed in the light of the potential use of these mitochondrial fractions in bioenergetic studies, or in the involvement of these organelles to stress response in conifers.

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Abbreviations

BSA:

Bovine serum albumin

DTE:

Dithioerythritol

EDTA:

Ethylenediaminetetraacetic acid

mΔΨ:

Mitochondrial electrical potential

PCD:

Programmed cell death

PEMs:

Pro-embryogenic cell masses

PVPP:

Polyvinylpolypyrrolidone

SE:

Somatic embryogenesis

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Acknowledgment

This research was supported by the University of Udine.

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Authors and Affiliations

  1. Department of Biology and Plant Protection, Section of Plant Biology, University of Udine, via Cotonificio 108, 33100, Udine, Italy

    Elisa Petrussa, Alberto Bertolini, Jana Krajňáková, Valentino Casolo, Francesco Macrì & Angelo Vianello

  2. National Forestry Center, Forest Research Institute, T. Masaryka 22, 96001, Zvolen, Slovakia

    Jana Krajňáková

Authors
  1. Elisa Petrussa
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  2. Alberto Bertolini
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  3. Jana Krajňáková
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  4. Valentino Casolo
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  5. Francesco Macrì
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  6. Angelo Vianello
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Corresponding author

Correspondence to Angelo Vianello.

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Communicated by L. Jouanin.

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Petrussa, E., Bertolini, A., Krajňáková, J. et al. Isolation of mitochondria from embryogenic cultures of Picea abies (L.) Karst. and Abies cephalonica Loud.: characterization of a K +ATP channel. Plant Cell Rep 27, 137–146 (2008). https://doi.org/10.1007/s00299-007-0436-2

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  • DOI: https://doi.org/10.1007/s00299-007-0436-2

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