Cryopreservation of Vitis vinifera via Droplet- vitrification | ||
| Arab Journal of Biotechnology | ||
| Volume 21, Issue 2, December 2018, Pages 147-158 PDF (2.43 M) | ||
| Document Type: Original Article | ||
| DOI: 10.21608/arjb.2018.428802 | ||
| Abstract | ||
| In the present study, two Egyptian grapevine cultivars (Red romy and Ghariby) and a Chinese variety (Cabernet sauvignon) were successfully cryopreserved by droplet vitrification. Axillary shoot tips were excised from two months old plantlets cultured on solidified ½MS medium with 0.5mg benzyladenine, 3% sucrose and 0.7% agar (pH 5.8) at 25 ◦C, under a 12 h light/12 h dark photoperiod with a light intensity of 40 μE m–2 s–1. For vitrification, excised shoot tips were precultured on half strength MS solidified medium supplemented with 0.1 M sucrose for 3 days in darkness and then treated with a mixture of 2 M glycerol and 0.4 M sucrose (LS solution) for 20 min at 25 ◦C. Shoot tips were then dehydrated with half-strength PVS2 vitrification solution (30% (w/v) glycerol, 15% (w/v) ethylene glycol, 15 % dimethylsulfoxide and 0.4 M sucrose in MS basal medium, for 30 min. This was followed by full strength PVS2 for (25min, 50 min or 60 min) at 0 ◦C before direct immersion in liquid nitrogen. The results showed that the mean percentage of survived shoot tips was not significantly different among the three genotypes, i.e., 66.67, 54.72 and 58.06% for Cabernet sauvignon, Red romy, and Ghariby, respectively. Also, the mean number of shoot tips regrowth was not significantly different, i.e. 57.50, 50.50 and 50.50 for Cabernet sauvignon, Red romy and Ghariby, respectively. The optimal duration of dehydration with PVS2 for survival and regrowth was 50 min. Ten ISSR primers were used for assessing the stability of the cryopreserved genotypes compared to the noncryopreserved. A negligible percentage of polymorphism was detected in the two cultivars Red romy (6.52%) and Ghariby (2.26%) with no morphological changes after cryopreservation. While, the cryopreserved plantlets of the cultivar Cabernet sauvignon did not exhibit any variability at the morphological or molecular levels compared to the control (noncryopreserved plantlets). | ||
| Keywords | ||
| Grapevine; cryopreservation; Droplet-vitrification; ISSR markers | ||
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