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以地被菊花(Dendranthema grandiflorum cv.White Snow)无菌苗叶片为外植体,在附加不同浓度植物生长调节剂的MS基础培养基上诱导培养,通过器官直接发生途径获得再生植株。结果表明,2.0mg/L6-BA和0.2mg/LNAA组合可获得93.8%的再生芽。将含拟南芥(Arabidopsis thaliana)逆境诱导转录因子DREB1A基因的植物表达载体pBI-DREB1A导入根癌农杆菌(Agrobacterium tumefaciens)LBA4404,农杆菌介导法遗传转化地被菊花White Snow叶盘,经PCR和PCR-Southern检测,DREB1A基因已整合到转化植株的基因组中。遗传转化过程中,预培养2d后,将OD600=0.5～0.7的根癌农杆菌稀释30倍后侵染叶盘10min,再共培养2d,有利于获得最高遗传转化效率。
Using sterile leaves of Dendranthema grandiflorum cv. White Snow as explants, cultured on MS basal medium supplemented with different concentrations of plant growth regulators, regenerated plants were obtained through organ-direct pathway. The results showed that 93.8% regenerated shoots could be obtained when the combination of 2.0mg / L 6-BA and 0.2mg / L NAA. The plant expression vector pBI-DREB1A containing the Arabidopsis thaliana stress-inducible transcription factor DREB1A gene was introduced into Agrobacterium tumefaciens LBA4404 and transformed into White Snow leaf discs by Agrobacterium tumefaciens mediated transformation by PCR And PCR-Southern assay, DREB1A gene has been integrated into the genome of the transformed plant. In the process of genetic transformation, the Agrobacterium tumefaciens with OD600 = 0.5-0.7 was inoculated 30 days after preincubation for 2 days and then infected with leaf discs for 10 minutes and co-cultured for 2 days, which was in favor of obtaining the highest genetic transformation efficiency.