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Cryopreservation of Pinus radiata embryogenic tissue: effects of cryoprotective pretreatments on maturation ability

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Abstract

The goal of the study was to quantify the effect of different cryoprotective pretreatments on the maturation ability of cryopreserved Pinus radiata embryogenic tissue. Fourteen cell lines were evaluated under five pretreatments: (1) Dimethyl sulfoxide 5% (v/v); (2) Dimethyl sulfoxide 10% (v/v); (3) Dimethyl sulfoxide 5% (v/v) and 0.09 M of l-proline; (4) Dimethyl sulfoxide 10% (v/v) and 0.09 M of l-proline; (5) 0.09 M of l-proline and compared to embryogenic tissue that was not cryopreserved (control). The cell lines were thawed at 37 °C to evaluate the recovery percentage and determine the maturation ability. The recovery rate of all cryopreserved material was significantly lower than the control and material treated with 0.09 M of l-proline, in absent dimethyl sulfoxide, showed the lowest levels of recovery. Regarding the number of embryos, an interaction between pretreatments and the cell lines under evaluation was identified. Moreover, morphological differences in the plants being treated with the most concentrated Dimethyl sulfoxide pretreatment was observed. At the microscopic level, no damage or morphological changes in proembryogenic masses were observed during the post-cryopreservation evaluation. Therefore, the best pretreatment was 5% Dimethyl sulfoxide supplemented with 0.09 M l-proline, allowing the most growth recovery of thawed samples and more cotyledonary embryos (maturation ability). The evaluated responses in presence of cryoprotective elements were genotype dependent.

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Abbreviations

ET:

Embryogenic tissue

Se:

Somatic embryo

DMSO:

Dimethyl sulfoxide

ECL:

Embryogenic cell line

ECLs:

Embryogenic cell lines

PEM:

Proembryogenic mass

PEMs:

Proembryogenic masses

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Acknowledgements

This study was carried out with the financing and plant material from the Forest research company Bioforest S.A. The authors thanks Dr. Mauricio Ramírez for his contribution on the reviewing of the manuscript and his collaboration in the statistical analysis.

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Authors and Affiliations

  1. Laboratorio de Cultivo de Tejidos Vegetales, Facultad de Ciencias Forestales y Centro de Biotecnología, Universidad de Concepción, Casilla 160-C, Concepción, Chile

    Yenny Lineros, Manuel Sánchez & Darcy Ríos

  2. Bioforest S.A., Unidad de Biotecnología, Camino a Coronel km 15 S/N, Coronel, Chile

    Yenny Lineros, Claudio Balocchi & Ximena Muñoz

Authors
  1. Yenny Lineros
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  2. Claudio Balocchi
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  3. Ximena Muñoz
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  4. Manuel Sánchez
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  5. Darcy Ríos
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Contributions

YL, DR performed the planning and design of experiments. YL performed the research and wrote the manuscript. CB XM helped with the technical support and review the results and MS read and approved the manuscript.

Corresponding authors

Correspondence to Yenny Lineros or Darcy Ríos.

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The author declares that they have no conflict of interest.

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Communicated by M. Angeles Revilla.

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Lineros, Y., Balocchi, C., Muñoz, X. et al. Cryopreservation of Pinus radiata embryogenic tissue: effects of cryoprotective pretreatments on maturation ability. Plant Cell Tiss Organ Cult 135, 357–366 (2018). https://doi.org/10.1007/s11240-018-1469-7

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  • DOI: https://doi.org/10.1007/s11240-018-1469-7

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