STANDARDIZING A STERILIZATION PROTOCOL FOR STEM NODAL CI]LTURE AND ESTABLISIIMENT OF IN VITRO MUXTIPLICATION RATES OF NEW TEA CULTWARS (Camellia sinensis (L.) O Kuntze)

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STANDARDIZING A STERILIZATION PROTOCOL FOR STEM NODAL CI]LTURE AND ESTABLISIIMENT OF IN VITRO MUXTIPLICATION RATES OF NEW TEA CULTWARS (Camellia sinensis (L.) O Kuntze) M.G.D. Malkanthir, K.K. Ranawe era,',M'A'B' Ranatunga', P.A. Weerasinghe' 'Department of Plant Sciences,Faculty of Agriculture, Raiarata University of Sri Lanka, Puliyankulama, Anuradhapurz, Sri Lanka' tplant Breeding Division, TbaResearchInstitute, Thlawakelle, Sri Lanka. of Successof tea micropropagationis hindered by severemicrobial contamination an at developing aims explants and low multiplication ratesoftea cultivars. The study measures effective sterilization protocol by combining pre harvesting sterilization six new from explants with five different surface sterilization protocols using nodal in tea tea cultivars. Attempts were also made to identiff most abundantcontaminants them. to control order in and to incorporate fungicide to the culture medium of Multiplication ratesof new cultivars were evaluatedunder different combinations (maintaining auxins and cytokinins. Impact on pre harvesting sterilization measures motherbushesinside apropagatorwith fungicide spraying)to reducecontaminations five surface of nodal explants was inconsistent among accessions.Among the yielded the sterilizationtechniquestested,treatmentwith 0.I%HgCl' for 10 minutes (45.25%). best results as it improved the number of clean and alive nodal explants were mixture 20 Treatments with acidified bleach and bleach, ethanol and tween bleach ineffective in reduction of contaminationse75%).Treatment with acidified resulted refrigerator a in and benlate mixture where cuttings were kept overnight severebrowning ofexplants(44.08%).Two fungiPestalotiopsisandAspergilluswere of the most prominent contaminantsthose resulted 65-70% and}SYocontaminations were explants respectively. When ftrngicides (Hexaconazole and Chlorothalonil) observed (>60%) were addedto the MS medium, high survival percentageof explants like HgCl' and that could be a viable alternative for hazardous surface sterilants preliminary results showed significant difference in multiplication rates among new rateaf 256 fold' tea cultivars and accession101 recordedthe highestmultiplication mgl-' of BAP (6Highestmultiplication ratewhich is 3. 12 fold was achievedwhen 2.5 usedin culture Benzyl-aminopurine)and 0.lmgl-'of IBA (Indole-3-butric acid) were medium. Key wordsz Camellia sinensls,Fungicide addedmedia, Multiplication explants,Pre steril izationmeasures

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rates,Nodal