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ThesisItem Open Access Effect of antibiotics on shoot regeneration and suppression of Agrobacterium growth in apple rootstock(NAUNI,SOLAN,UHF, 2011) KASHYAP, MONIKA; MODGIL, MANJUABSTRACT The present investigation aims at “Studies on cell culture for genetic transformation in Punica granatum L.”. Genetic transformation experiment was conducted with cotyledonary explants excised from in vitro raised seedlings of Punica granatum L. cv. Kandhari Kabuli. Explants were preconditioned on standardized callus induction medium (solid MS medium supplemented with 4.0 mg/l BA and 3.0 mg/l NAA) for 48 hours followed by 48 hours of cocultivation with Agrobacterium tumefaciens. Friable putative transgenic were used for initiation of cell suspension culture. Cells of various shapes such as oval, elongated, columnar, elliptical and isodiametric were observed in the suspension culture. Cells proliferated best on selective MS medium supplemented with 4.0 mg/l BA + 3.0 mg/l NAA and 50 mg/l kanamycin along with 500 mg/l cefotaxime with average cell doubling time of 5 days. Combination of a cytokinin (BA) and an auxin (NAA) was found to be better for cell proliferation as compared with the combination of two cytokinins (BA and TDZ). Growth curve of cells in suspension culture consisted of lag phase (5-6 days), exponential phase (7 days) and stationary phase (4-5 days) followed by cell death. Plating efficiency of 0.7 % was obtained by plating of cell aggregates (5-6 cells) on selective MS medium supplemented with 4.0 mg/l BA and 3.0 mg/l NAA utilizing modified Bergman technique. Putative transgenic macrocalli did not show shoot regeneration. Positive PCR results were observed in transgenic calli obtained through cell suspension culture. The protocol, thus offer an opportunity to produce the putative transformed tissues/ plantlets with single cell origin.