【摘要】 设计引物并克隆了烟草叶绿体16SrDNA基因的启动子,并在其下游加入了rbcL基因的SD序列,以使该启动子能有效翻译外源基因;进一步以aadA基因作为外源基因,以psbA基因的3’序列为终止子,构建成aadA基因表达盒;将此表达盒插入烟草叶绿体同源片段trnK-psbA-trnH中,构建成烟草叶绿体转化及表达载体P16T.用基因枪微弹轰击转化法,将该载体导人野生型大烟草叶绿体中,建立了烟草叶绿体遗传转化体系,转化频率为0.5株/枪.对转化植株进行PCR及遗传杂交实验分析,证明外源基因已在烟草叶绿体中稳定整合并遵循母系遗传规律.更多还原

【Abstract】 A Nicotiana tabacum chloroplast transformation vector p16T has been constructed on the basis of a mutated 16S rDNA promoter. It contains not only aadA. gene regulated by 16S promoter and psbA teminator, but also two homologous fragments, trnH-psbA and trnK. Using biolistic method, the plasmid p16T has been transformed into the wild type Nicotiana tabacum leaves. Through spectinomycin selection, transformed calli and regenerated plants were obtained from tobacco seedling leaves. The result of PCR analysis showed that the foreign gene had been transformed into the chloroplast of tobacco. Hybrid seedlings of transgenic tobacco acting as the female parent showed the resistance to spectinomycin with the mode of maternal heredity.更多还原