In vitro studies on morphogenesis and plant regeneration in elite clones of cocoa (theobroma cacao L.)
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Date
2018
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Department of Plantation Crops and Spices College of Horticulture, Vellanikkara
Abstract
The experiment entitled “In vitro studies on morphogenesis and plant regeneration in elite clones of cocoa (Theobroma cacao L.) was carried out in Tissue culture laboratory of Department of Plantation Crops and Spices, College of Horticulture, Vellanikkara during 2016-2018 with an objective to develop a protocol for in vitro regeneration and also to determine the variation in morphogenesis and regeneration among elite cocoa clones. The experiment was conducted using five varieties, CCRP 2, CCRP 6, CCRP 8, CCRP 15 and Scavina. Single nodded cuttings of 2-3 cm collected from the budded plants were used as explants. The best treatment for surface sterilization of nodal segments was washing in Teepol followed by shaking in 0.2 per cent Mancozeb for 30 minutes and then, Streptocycline 0.1 per cent for 10 minutes outside the laminar air flow chamber. Inside the laminar air flow chamber, the explants were treated with Ethyl Alcohol 70 per cent for 3 minutes followed by HgCl2 0.1 per cent for 5 minutes. This treatment showed the highest survival percentage (91.67%) of cultures with minimum microbial contamination (8.33%). The medium WPM + 2-ip 1ppm + IAA 0.02ppm showed cent per cent shoot induction in varieties CCRP 15 and Scavina; WPM + 2-ip 2ppm + IAA 0.04ppm in varieties CCRP 2, CCRP 6 and Scavina; WPM + 2-ip 3ppm + IAA 0.06ppm in CCRP 8 and WPM + 2-ip 4ppm + IAA 0.08ppm in varieties CCRP 2 and Scavina. Period of shoot induction for all the five varieties was significantly influenced by media, varieties and their interactions. Among the media, the lowest period of shoot induction was recorded in medium WPM + 2-ip 1ppm + IAA 0.02ppm (8.41). Among the varieties, the lowest period of shoot induction was in CCRP 2 (7.10) and this was statistically on par with variety Scavina (8.37). Among the interactions, the period of shoot induction was lowest in variety CCRP 15 in medium WPM + 2-ip 1ppm + IAA 0.02ppm (6.33). Culture establishment percentage in multiple shoot induction media varied significantly among media. Highest culture establishment percentage (95%) was recorded in media WPM + 2-ip 3ppm + IAA 0.06ppm + AgNO3 5ppm and WPM + 2- ip 4ppm + IAA 0.08ppm + AgNO3 5ppm. Among the varieties, highest multiple shoot induction percentage was recorded in Scavina (93.75%). Interaction effect of media and varieties on number of multiple shoots per culture was significantly different and the highest number of multiple shoots per culture (2.25) was observed in variety Scavina when medium WPM + 2-ip 4ppm + IAA 0.08ppm + AgNO3 5ppm was used. Rooting percentage was generally low under in vitro condition. But among the media tried, 1⁄2 MS + IBA 4000 ppm (pulse treatment for 2-3 sec) and 1⁄2 MS + IBA 5000 ppm (pulse treatment for 2-3 sec) were found to be ideal. All the varieties showed root induction in 1⁄2 MS + IBA 5000 ppm (pulse treatment for 2-3 sec) and only two varieties, CCRP 15 and Scavina, had successful root induction in 1⁄2 MS + IBA 4000 ppm (pulse treatment for 2-3 sec). Hence in general, it can be concluded that, 1⁄2 MS + IBA 5000 ppm (pulse treatment for 2-3 sec) was superior to all the media tried for root induction. Hardening of tissue culture cocoa plants were not successful in both sand: vermicompost (1:1) and vermiculite: perlite: vermicompost (1:1:1). The vermiculite: perlite: vermicompost (1:1:1) media showed a maximum survival of plantlets for one week. However, later showed yellowing and finally the shoots were completely dried off. The anatomical observation on root-shoot transition zone using rotary microtome sectioning showed the presence of intervening callus between the vascular tissues of root and shoot. This callus inhibits proper vascular connection and thus, the plants kept for hardening were difficult to get established.
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Master of Science in Horticulture
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