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Baskar, Venkidasamy; Gangadhar, Baniekal H; Park, Se Won; Nile, Shivraj Hariram
3 Biotech, 06/2016, Volume: 6, Issue: 1Journal Article
The present study aims to investigate the numerous factors influencing Agrobacterium tumefaciens-mediated genetic transformation of Chinese cabbage (Brassica rapa ssp. pekinensis). Factors affecting transformation efficiency, such as age of explants, Agrobacterium concentration, and effect of acetosyringone, pre-cultivation, infection and co-cultivation time of Agrobacterium were examined. The pre-cultured hypocotyls from young seedlings prior to exposure to Agrobacterium showed higher shoot regeneration. The plant transformation with the modest A. tumefaciens concentrations (0.8 OD) and the 3 days co-cultivation periods increased transformation efficiency. Plant growth hormones 1-naphthyl acetic acid (NAA) and 6-benzyl amino purine (BAP) were essential for callus and shoot formation. Root formation was effective in half strength MS medium without supplementation of root-inducing hormones. To maintain selection pressure, plant subculture was carried out every 2 weeks with selective antibiotics. The putative transgenic plants were acclimatized in the greenhouse. Polymerase chain reaction was performed to confirm the integration of T-DNA into the genome of transgenic plants. A transformation efficiency of 15 % was obtained. This protocol allows effective transformation and indirect regeneration of Brassica rapa.
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