Studies on tissue culture and Agrobacterium-mediated transformation system of Chrysanthemum

• Main Authors: Li-Yu Wu, 吳麗玉
• Other Authors: Li-Jen Liao
• Format: Others
• Language: zh-TW
• Published: 2005
• Online Access: http://ndltd.ncl.edu.tw/handle/78015771781410182363

碩士 === 國立高雄師範大學 === 生物科學研究所 === 93 === Using gene transformation technology for inserting foreign target gene(s) into plant chromosome, following by tissue culture techniques for regeneration of transgenic plants, however the regeneration protocols were developed mostly only suitable for the specific cultivar(s) and unable to apply to the numerous varieties. In this study, regeneration systems using 〝Huang Jing Ching〞cultivars of chrysanthemum were established. Using petal explants do tissue culture. The best rate of somatic embryo and bud formation from explants were cultured on the MS medium containing 5 mg/L of IAA, 5 mg/L of BA and 0.05 mg/L of Kinetin. The aseptic leaf discs and internodes were chosen as the explants of Agrobacterium-mediated transformation. Using Agrobacterium strain EHA105 and LBA4404-mediated transformation system. Cefotaxime were tested for bacteriostatic effects and influences on regeneration. It was found that Cefotaxime had obvious suppressing influence on shoots regeneration of the chrysanthemum. A study of proper Hygromycin B concentration for screening putative transgenic plants was also conducted, the 10 mg/L Hygromycin B was found effective for shoot tip growthing. Using GUS and Hygromycin B as a early selection method for the putative transgenic plants. Putative transgenic plants were further analyzed by PCR amplification. This study obtain 8 putative lines of HSF transgenic plants, and 7 putative lines of IPT transgenic plants . Further investigations for the HSF and IPT gene expression and improving of Agrobacterium-mediated transformation system for chrysanthemum are needed for the future work.