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Vitrification and a heat-shock treatment improve cryopreservation of tobacco cell suspensions compared to two-step freezing

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Abstract

Vitrification and two-step freezing were comparatively tested for cryopreservation of tobacco cell suspensions. The optimal growth phase of the culture and the optimal length of the protecting preculture period were determined. With both methods, late-exponential cells showed higher survival rates compared to early exponential and growth-limited cells. Under optimal conditions vitrification yielded higher survival rates than two-step freezing (55% and 36%, respectively). Using two-step freezing a preculture period of 72 h in medium supplemented with 0.3 M mannitol was necessary to obtain maximal survival, whereas for vitrification 24 h of preculture sufficed. Heat shock treatment prior to the cryopreservation procedure could improve survival when mannitol precultured cells in a non-optimal growth phase were used. Heat-shocked cells, which were not precultured with mannitol, did not survive vitrification. Vitrification is the method recommended for cryopreservation of tobacco cell suspensions, in view of the shorter preculture period and higher survival rates resulting in quicker regrowth.

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Authors and Affiliations

  1. Institute of Molecular Plant Sciences, Clusius Laboratory, Leiden University, Wassenaarseweg 64, 2333 AL, Leiden, The Netherlands

    Poula J. Reinhoud, Elly W. M. Schrijnemakers, Frank van Iren & Jan W. Kijne

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  1. Poula J. Reinhoud
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  2. Elly W. M. Schrijnemakers
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  3. Frank van Iren
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  4. Jan W. Kijne
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Reinhoud, P.J., Schrijnemakers, E.W.M., van Iren, F. et al. Vitrification and a heat-shock treatment improve cryopreservation of tobacco cell suspensions compared to two-step freezing. Plant Cell Tiss Organ Cult 42, 261–267 (1995). https://doi.org/10.1007/BF00029997

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