Keywords
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callus induction, Citrus grandis, cotyledon, pummel, shoot regeneration
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Abstract
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Pummelo, a citrus fruit, is an underutilized fruit crop with a potential for
commercialization in warm humid climates. Improvement of crops through biotechnological
approaches mainly involves availability of efficient in vitro regeneration
protocols. A study was conducted to develop a regeneration protocol for a local
cultivar of pummelo under cultivation in Devanahalli, Bangalore, India, using
cotyledon explants obtained from six-week-old in vitro grown seedlings. Callus was
induced on MS (Murashige and Skoog) media by use of α-naphthaleneacetic acid
(NAA) at 1, 3 and 5 mg/L, and 2,4-dichlorophenoxyacetic acid (2,4-D) at 0.5, 1 and 2
mg/L, each one individually and also in combination with 6-benzyl adenine (BA) at
concentrations of 0.5 and 1 mg/L. The best callusing was obtained in media containing
5 mg/L NAA plus 0.5 mg/L BA (100% callusing with 1.2 g callus fresh weight per
explant). In order to induce shoots in the calli, they were transferred in MS media
containing each of BA (1 to 8 mg/L) and Adenine Sulphate (AdS) (1 to 40 mg/L), with
or without 0.25 mg/L indole-3-butyric acid (IBA). 2,4-D-induced calli showed ability of
regenerating shoots better than NAA-induced calli which failed almost totally. Shoot
regeneration was best (90% with average 1.9 shoots/explant) when BA was used at 5
mg/L alone. AdS was less effective than BA in shoot induction in the calli though its
best result, 70% shoot induction with average of 2 shoots/explant at 20 mg/L, was also
considerable. Presence of IBA in the media with each of the two cytokinins used,
generally, affected shoot regeneration negatively. The regenerated microshoots were
rooted in half strength MS medium supplemented with 4 mg/L IBA. The produced
protocol in this study can be useful for improvement of pummelo through
biotechnological techniques which involve callus tissue.
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