Lekha Rani, CRahana, S NKAU2019-06-222019-06-222006http://krishikosh.egranth.ac.in/handle/1/5810109630PGAttempts were made to undertake initial culture establishment and molecular characterization of 40 selected Dendrobium hybrids developed in the Department of Plant Breeding and Genetics under a DBT funded orchid breeding project and to refine the protocol for rapid multiplication using in vitro leaf explants. The studies were carried out in the Department of Plant Breeding and Genetics, College of Agriculture, Vellayani during 2004-2006. Initial culture establishment of the 40 selected Dendrobium hybrids were carried out using stem nodal explants obtained from keikis and were inoculated in the identified best media, viz., VW + KN 4 mg l-1 + IAA 4 mg l-1 + CW 200 ml l-1 (Sivamani, 2004). The nature of response of the stem nodal explant was found to be varied among the hybrids. The duration taken for the initiation of PLBs, shoot, leaf and root differed significantly among the hybrids. Once the PLBs or shoot buds had been differentiated, the cultures were transferred to the subculturing media for faster growth and development. Leaves and roots appeared from the shoot buds or PLBs after four to ten weeks in culture. The plantlets obtained were hardened and transferred to the greenhouse. Plantlets obtained through stem nodal cultures were more vigorous than that from in vitro leaf explant cultures. The regeneration potential of in vitro sourced leaf explants appeared to be an inherent trait of orchids which gets evoked under certain physico-chemical stimulus under in vitro condition. The in vitro sourced leaf explants can be profitably used for micropropagation. The main advantage of this technique is that in selected single plant hybrids where keiki production is low, once they are established in vitro, they can be further mass multiplied using the in vitro leaf as explant. The young leaf segments derived from in vitro established Dendrobium hybrids were used as explant. Swelling of cut leaf edges and callus initiation studies were conducted with two levels of CH (500 mg l-1 and 250 mg l-1) in half strength MS medium containing BAP 5 mg l-1 + KN 5 mg l-1 + CH 500 mg l-1 + CW 200 ml l-1 + AC l gl-1. Out of this CH at 500 mg l-1 was found to be the best for callus initiation and subsequent development of plantlet. The callus obtained from the inoculation medium, irrespective of the levels of CH used, were grouped together and cultured on regeneration media with two different levels of CH, viz., CH at 250 mg l-1 and no CH. Presence of CH at 250 mg l-1 was found to have significant effect on shooting response. The optimum medium combination for shoot initiation was found to be half strength MS medium with BAP 5 mg l-1 + KN 5 mg l-1 + CH 250 mg l-1 + CW 200 ml l-1 + AC 1 gl-1. Out of the different strengths of MS medium tried with in vitro leaf explant, half strength MS medium exhibited early establishment of plantlet and recorded minimum number of days for deflasking. Half strength MS with NAA 0.1 mg l-1 + BAP 0.5 mg l-1 + AC 1 gl-1 + CW 200 mg l-1 was found to be the best medium combination for better in vitro rooting and early establishment of plantlet. Plantlets were ready for plant out in six months. Random amplified polymorphic DNA (RAPD) analysis was employed for the molecular characterization of the 40 selected Dendrobium hybrids. The eight primers, selected from the 40 initially screened primers, generated 69 scorable bands of which three were monomorphic and the remaining 66 were polymorphic (95.65 %). All primers produced polymorphic amplification products, however, the extent of polymorphism varied with each primer. Statistical analysis was carried out using NTSYS-PC software and a dendrogram was generated using Jaccard’s similarity coefficients. The overall similarity coefficient ranged from 0.29 to 1.00. The forty selected Dendrobium hybrids that were studied formed 11 clusters in UPGMA cluster analysis. The grouping of hybrids in the cluster was largely consistent with what is known about their breeding history. The cluster based on RAPD analysis using eight primers clearly demonstrates the existence of genetic variation within the 40 selected Dendrobium hybrids. Polymorphism obtained in the present study can be used as fingerprints of the forty selected Dendrobium hybrids.ennullThesis