根癌农杆菌介导的花魔芋遗传转化体系的研究

本研究以潮霉素抗性基因为选择标记,建立了根癌农杆菌介导的花魔芋(Amorphophalluskonjac)的遗传转化体系。同时探讨了魔芋愈伤组织的培养基、筛选剂和筛选时间等因素对转化效率的影响。通过筛选时间不同的两个转化试验,分别从300个花魔芋愈伤组织中得到了16个和18个具有潮霉素抗性的愈伤组织,其抗性愈伤率为5.33%和6.00%。获得的抗性愈伤组织进一步在分化培养基上进行分化,其分化效率分别为31.2%和22.2%。移栽后,分别得到了39株和36株成活植株。PCR检测表明分别有26株和21株再生花魔芋出现了800bp的特异性扩增带型,其阳性率分别为66.7%和58.3%。随机挑取20个PCR阳性植株的DNA进行斑点杂交检测,结果表明外源基因已经整合到花魔芋的基因组中。

Genetic Transformation System of Amorphophallus konjac Mediated by A grobacterium tumfaciens

An Agrobacterium tumfaciens-mediated transformation system has been developed for the production of transgenic plants of Amorphophallus konjac. The explants were inoculated with an Agrobacterium tumfaciens strain EHA105 carrying the plasmid pBMB0890 containing selective gene hygromycin phosphotransferase (hpt). Factors that influence Agrobacterium tumfaciens-mediated transformation of Amorphophallus konjac, including culture medium (Table 1), selection agents and selection time were investigated. 16 and 18 resistant calli were obtained from 300 calli separately. The transformation frequency was 5.33% and 6.00%, repectively. 31.2% and 22.2% of resistant calli could generate plants. PCR analysis of the 39 and 36 live plants indicated 26 and 21 plants were integrated with foreign DNA separately, resulting 66.7% and 58.3% transgenic ratio, respectively. 20 plants of those transgenic plants were re-clarified by spot blot hybridization.