Plant Regeneration Through Callus Organogenesis and True-to-Type Conformity of Plants by RAPD Analysis in Desmodium gang
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Plant Regeneration Through Callus Organogenesis and True-to-Type Conformity of Plants by RAPD Analysis in Desmodium gangeticum (Linn.) DC. Meena K. Cheruvathur & Jyothi Abraham & T. Dennis Thomas
Received: 20 August 2012 / Accepted: 14 January 2013 / Published online: 23 January 2013 # Springer Science+Business Media New York 2013
Abstract An efficient plant regeneration protocol was established for an endangered ethnomedicinal plant Desmodium gangeticum (Linn.) DC. Morphogenic calli were produced from 96 % of the cultures comprising the immature leaf explants on Murashige and Skoog (MS) medium supplemented with 2,4-dichlorophenoxyacetic acid (4.0 mgl−1) in combination with 6benzylaminopurine (BA; 0.8 mgl−1). For callus regeneration, various concentrations of BA (1.0–5.0 mgl−1) or thidiazuron (TDZ; 1.0–5.0 mgl−1) alone or in combination with indole-3acetic acid (IAA; 0.2–1.0 mgl−1) were used. Highest response of shoot regeneration was observed on MS medium fortified with TDZ (4.0 mgl−1) and IAA (0.5 mgl−1) combination. Here, 100 % cultures responded with an average number of 22.3 shoots per gram calli. Inclusion of indole-3-butyric acid in half MS medium favored rooting of recovered shoots. Out of 45 rooted plants transferred to soil, 40 survived. Total DNA was extracted from the leaves of the acclimatized plants of D. gangeticum. Analysis of random amplified polymorphic DNA using 13 arbitrary decanucleotide primers showed the genetic homogeneity in all the ten plants regenerated from callus with parental plant, suggesting that shoot regeneration from callus could be used for the true-to-type multiplication of this plant. Keywords Callus . Desmodium gangeticum . Immature leaf . RAPD analysis . Regeneration Abbreviations BA N6-Benzylaminopurine 2,4-D 2,4-Dichlorophenoxyacetic acid IAA Indole-3-acetic acid IBA Indole-3-butyric acid MS Murashige and Skoog (1962) NAA 1-Naphthalene acetic acid RAPD Random amplified polymorphic DNA TDZ Thidiazuron M. K. Cheruvathur : J. Abraham : T. D. Thomas (*) Postgraduate and Research Department of Botany, St. Thomas College, Pala, Arunapuram (P.O.), 686574 Kottayam (Dt.), Kerala, India e-mail: [email protected]
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Appl Biochem Biotechnol (2013) 169:1799–1810
Introduction Desmodium gangeticum (Linn.) DC. is an endangered ethnomedicinal plant belongs to Fabaceae family [1]. It is an erect woody undershrub in tropical region with alternate leaves and compressed fruits. This plant is one among the Dashamoola (ten roots) of Ayurveda and is an important ingredient of many famous Ayurvedic drugs like Dashamoolarishta, Chyavanaprasha, Dhanwantharam Kuzhambu, Rasnadhi decoction, and Brahma Rasayan [2, 3]. D. gangeticum is of great therapeutic value in treating diseases such as typhoid, piles, inflammation, asthma, and bronchitis [4]. The plant is mineral rich, and the whole plant decoction is given to treat digestive disorders, edema, diarrhea, intermittent fevers, malaria, and urinary tract infections [5]. In folklore medicine, decoction from D. gangeticum leaves used for ston
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