Regeneration optimisation of cucumber (Cucumis sativus L.) and GFP (green fluorescent protein) gene transformation via agrobacterium tumefaciens aler

ABSTRACTMSc THESISREGENERATION OPTIMISATION OF CUCUMBER (Cucumissativus L. ) and GFP (GREEN FLUORESCENT PROTEIN) GENETRANSFORMATION via Agrobacterium tumefaciensSinem SERBEST-KOBANERDEPARTMENT OF BIOTECHNOLOGYINSTITUTE OF NATURAL AND APPLIED SCIENSESUNIVERSITY OF CUKUROVAFirst Supervisor : Doç.Dr. Neslihan Yeşim YALÇIN-MENDİYear : 2006, Page: 97Jury : Doç.Dr. Neslihan Yeşim YALÇIN-MENDİProf.Dr. Selim ÇETİNERDoç.Dr. Mukaddes KAYIMAn efficient method for shoot regeneration and Agrobacterium mediated genetransformation of Cucumber (Cucumis sativus L.) has been developed in this study.The optimum regeneration and transformation protocol were tested. The use ofcotyledon explants excised from 4 day-old seedlings cultured in Murashige andSkoog?s medium, supplemented with 1mg/l BAP and 1 mg/l IAA, resulted in a highregeneration frequency (% 82). Regeneration protocol has been successfully appliedto obtain transformed plants, using Agrobacterium tumefaciens strain AL-1 which isharboring a binary vector pBin m-GFP5-ER carrying npt II (selectable marker gene)and mGFP5-ER (Haseloff, 1995) (reporter gene), encoding green fluorescent protein,driven by cauliflower mosaic virus 35S promoter. Cotyledon explants, infected byAgrobacterium suspansion, cocultivated in the MS medium supplemented with 2mg/l BAP, 1 mg/l IAA, 200 μM acetosyringon, 0.5 g/l MES and phytagel.Transformed tissues were detected at a very earyl stage at 254/365nm.emmition byhand UV.Keywords: Cucumber, regeneration, mGFP5-ER, transformation, AgrobacteriumtumefaciensII