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Cryopreservation of in vitro-grown shoot tips of strawberry by the vitrification method using aluminium cryo-plates

  • Shin-ichi Yamamoto (a1), Kuniaki Fukui (a1), Tariq Rafique (a2), Nayyar Iqbal Khan (a3), Carlos Roman Castillo Martinez (a4), Kentaro Sekizawa (a5), Toshikazu Matsumoto (a6) and Takao Niino (a1)
  • DOI:
  • Published online: 28 November 2011

Cryopreservation using an aluminium cryo-plate was successfully applied to in vitro-grown strawberry (Fragaria × ananassa Duch.) shoot tips. The shoots were cold-hardened at 5°C for 3 weeks with an 8-h photoperiod. The shoot tips (1.5–2.0 mm × 0.5–1.0 mm) were dissected from the shoot and pre-cultured at 5°C for 2 d on Murashige and Skoog medium containing 2 M glycerol and 0.3 M sucrose. The pre-cultured shoot tips were placed on the aluminium cryo-plate containing ten wells embedded in alginate gel. Osmoprotection was performed by immersing the cryo-plates in a loading solution (2 M glycerol and 0.8 M sucrose) for 30 min at 25°C. Dehydration was performed by immersing the cryo-plates in plant vitrification solution 2 for 50 min at 25°C. Then, the cryo-plate with shoot tips was transferred into an uncapped cryotube that was held on a cryo-cane and directly immersed into liquid nitrogen (LN). After storage in LN, shoot tips attached to the cryo-plate were directly immersed into 2 ml of a 1 M sucrose solution for regeneration. Using this procedure, the average regrowth level of vitrified shoot tips of 15 strawberry cultivars reached 81%. This new method has many advantages and will facilitate the cryostorage of strawberry germplasm.

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D Hirai and A Sakai (1999) Cryopreservation of in vitro-grown meristems of potato (Solanum tuberosum L.) by encapsulation–vitrification. Potato Research 42: 153160.

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D Hirai , K Shirai , S Shirai and A Sakai (1998) Cryopreservation of in vitro-grown meristems of strawberry (Fragaria × ananassa Duch.) by encapsulation–vitrification. Euphytica 101: 109115.

T Murashige and F Skoog (1962) A revised medium for rapid growth and bioassays with tobacco tissue cultures. Physiolgia Plantarum 15: 473479.

T Niino , D Tanaka , S Ichikawa , J Takano , S Ivette , K Shirata and M Uemura (2003) Cryopreservation of in vitro-grown apical shoot tips of strawberry by vitrification. Plant Biotechnology 20: 7580.

A Sakai , S Kobayashi and I Oiyama (1990) Cryopreservation of nucellar cells of navel orange (Citrus sinensis Osb. Var. brasiliensis Tanaka) by vitrification. Plant Cell Reports 9: 3033.

A Sakai , D Hirai and T Niino (2008) Development of PVS-based vitrification and encapsulation–vitrification protocols. In: B Reed (ed.) Plant Cryopreservation: A Practical Guide. New York: Springer LLC, pp. 3358.

K Sekizawa , S Yamamoto , T Rafique , K Fukui and T Niino (2011) Cryopreservation of in vitro-grown shoot tips of carnation (Dianthus caryophyllus L.) by vitrification method using aluminium cryo-plates. Plant Biotechnology 28: 401405.

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Plant Genetic Resources
  • ISSN: 1479-2621
  • EISSN: 1479-263X
  • URL: /core/journals/plant-genetic-resources
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