Abstract

Promoters are key elements regulating gene expression levels, therefore their selection is an important step in genetic engineering research. The reporter gene gfp, which encodes green fluorescent protein (GFP), was transiently expressed in leaf tissues of Aztec tobacco Nicotiana rustica L. Compared to other species of the Nicotiana genus, Aztec tobacco has a large potential for expression of heterologous proteins, a large vegetative biomass, can be easily infiltrated, and is unpretentious in cultivation. Six genetic constructs were used with different promoter sequences: the 35S promoter of Cauliflower Mosaic Virus (35S CaMV), the double-enhanced 35S promoter (D35S CaMV), promoters of the RbcS1B and RbcS2B genes encoding the small subunit of ribulose-1,5-bisphosphate carboxylase/oxygenase (Rubisco) isolated from Arabidopsis thaliana (L.) Heynh., and promoters of the LHB1B1 and LHB1B2 genes from A. thaliana encoding chlorophyll a/b binding proteins. The gfp gene expression was detected visually, spectrofluorimetrically, and by protein content (Bradford assay) on the seventh day after infiltration. The highest level of expression was observed using the double-enhanced 35S promoter (D35S CaMV) and the lowest using the LHB1B1 gene promoter.

中文翻译：

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不同启动子序列构建的农杆菌介导的烟草瞬时转化后gfp基因表达水平的比较

摘要

启动子是调节基因表达水平的关键元素，因此，启动子的选择是基因工程研究的重要步骤。编码绿色荧光蛋白（GFP）的报告基因gfp在阿兹台克烟草Nicotiana Rusta L的叶片组织中瞬时表达。与其他烟草属相比，阿兹台克烟草具有很大的表达异源蛋白的潜力，营养生物质，易于渗透，在种植中没有伪装。使用了六个具有不同启动子序列的基因构建体：花椰菜花叶病毒的35S启动子（35S CaMV），双重增强的35S启动子（D35S CaMV），RbcS1B和RbcS2B的启动子拟南芥（L.）Heynh。分离的编码核糖-1,5-双磷酸羧化酶/加氧酶（Rubisco）小亚基的基因，以及拟南芥编码叶绿素a / b结合蛋白的LHB1B1和LHB1B2基因的启动子。所述GFP目视检测基因表达，spectrofluorimetrically，并通过在浸润后第7天的蛋白质含量（Bradford测定）。使用双重增强的35S启动子（D35S CaMV）观察到最高的表达水平，而使用LHB1B1基因启动子观察到最低的表达水平。