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本研究利用转基因烟草分析了mi R156对SPL3基因的保守性调节作用。首先通过数据库搜索和RT-PCR方法克隆了普通烟草Ntab SPL3基因的编码序列,并进行了测序验证。同时从拟南芥Col-0生态型基因组DNA分离了At Pri-mi R156a基因序列,构建产生植物表达载体,采用农杆菌转化技术制备了转基因烟草植株。RT-PCR分析发现,过量表达Atmi R156a可以明显下调烟草Ntab SPL3基因。表型观察结果显示,At Pri-mi R156a转基因烟草个体矮小,开花时间明显延迟,叶片数量及生物量积累增多,说明组成性表达Atmi R156a能够延长普通烟草的营养生长时间,推迟开花转变过程。本研究为培育适合不同生长环境的烟草新品种提供了一条新的途径,对烟草改良具有重要意义。
In this study, the conserved regulatory effect of mi R156 on SPL3 gene was analyzed using transgenic tobacco. Firstly, the coding sequence of Ntab SPL3 gene of Nicotiana tabacum was cloned by database search and RT-PCR method and sequenced. At the same time, At Pri-mi R156a gene was isolated from the genomic DNA of Col-0 in Arabidopsis thaliana, and the plant expression vector was constructed. The transgenic tobacco plants were prepared by Agrobacterium tumefaciens transformation. RT-PCR analysis showed that Overexpression of Atmi R156a significantly down-regulated tobacco Ntab SPL3 gene. Phenotypic observation showed that At Pri-mi R156a transgenic tobacco plants were short, the flowering time was significantly delayed and the number of leaves and biomass accumulation increased, indicating that the constitutive expression of Atmi R156a can prolong the vegetative growth of tobacco and delay the flowering process. This study provides a new way to cultivate new varieties of tobacco suitable for different growth environments, which is of great significance for tobacco improvement.