, Volume 255, Issue 4, pp 1065–1077 | Cite as

Cold, antioxidant and osmotic pre-treatments maintain the structural integrity of meristematic cells and improve plant regeneration in cryopreserved kiwifruit shoot tips

  • Liya Mathew
  • Andrew McLachlan
  • Rubina Jibran
  • David J. Burritt
  • Ranjith PathiranaEmail author
Original Article


Cryopreservation is a reliable and cost-effective method for the long-term preservation of clonally propagated species. The number of vegetatively propagated species conserved by cryopreservation is increasing through development of vitrification-based methods; droplet vitrification in particular is becoming the preferred method for many species, as it ensures fast freezing and thawing rates. This research investigated if cold, antioxidant and osmotic pre-treatments could maintain the structural integrity of cells, thence aid in developing a droplet vitrification protocol for kiwifruit using Actinidia chinensis var. chinensis ‘Hort16A’ as a model. Cold acclimation of donor plantlets at 4 °C for 2 weeks followed by sucrose pre-culture of shoot tips and supplementation of ascorbic acid (0.4 mM) in all media throughout the procedure registered 40% regeneration after cryopreservation. Transmission electron microscope imaging of meristematic cells confirmed sucrose and ascorbic acid pre-treatment of shoot tips from cold acclimated plantlets following treatment in vitrification solution exhibited severe plasmolysis and some disruption of membrane and vacuoles. In contrast cells without cold acclimation or sucrose and ascorbic acid pre-treatments exhibited minimal change after exposure to vitrification solution. After cryopreservation and recovery, all cells of untreated shoot tips showed rupture of the plasma membrane, loss of cytoplasmic contents and organelle distortions. By comparison, most pre-treated shoot-tip cells from cold acclimated plantlets retained their structural integrity, showing that only those cells that have been dehydrated and plasmolysed can withstand cryopreservation by vitrification.


Actinidia chinensis Ascorbic acid Droplet vitrification Tissue culture Sucrose Transmission electron microscopy Ultrastructure 



The authors thank Andrew Mullan and Belinda Diepenheim for the technical assistance, Tony Corbett for the preparation of figures, and Dr. Jayanthi Nadarajan for the critical reading of the manuscript.

Compliance with ethical standards

Conflicts of interest

The authors declare that they have no conflicts of interests.

Supplementary material

709_2018_1215_MOESM1_ESM.pdf (238 kb)
Online Resource 1 (PDF 237 kb)


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Copyright information

© Springer-Verlag GmbH Austria, part of Springer Nature 2018

Authors and Affiliations

  1. 1.The New Zealand Institute for Plant and Food Research LimitedPalmerston NorthNew Zealand
  2. 2.Department of BotanyUniversity of OtagoDunedinNew Zealand

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