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高活性启动子在基因时空表达调控方面有着重要的作用,利用高效率启动子调控抗性目的基因特异性表达,不仅可以提高作物抗胁迫能力,而且对改良品种方面具有重要的意义。迄今为止,有关水稻翻译起始因子GOS2基因(eukaryotic translation initiation factor 1b)启动子p GOS2启动效率鉴定的报道还很有限,尤其是与Ca MV35S双拷贝启动子的启动效率评估的研究还未见报道。本研究旨在利用PCR和生物信息学等技术,分离及解析水稻GOS2启动子特性,构建p GOS2::GUS双元表达载体,使用农杆菌介导法转化水稻胚性愈伤组织并获得转基因水稻植株,并通过PCR检测与GUS组织化学分析,评估p GOS2在转基因水稻叶片组织中的效率。生物信息学分析结果表明:所克隆的GOS2启动子,序列全长为3 155 bp,具有真核生物典型的一些顺式元件,如TATA-box、AGA-motif、CAAT-box以及GCGC-repeat等;GOS2启动子核心序列位于-43 bp~+4 bp区,得分是0.97。GUS组织化学分析结果显示:阴性对照组未发现GUS信号,而转基因实验组则表现为不同程度的GUS活性,p GOS2启动效率远远高于单双拷贝的Ca MV35S启动子。本研究结果证实了GOS2启动子在水稻叶片组织中的活性远远地高于加强型Ca MV35S启动子,为今后水稻分子育种及相关启动子的开发与筛选提供了一种可行的方法和新的视野。
Highly active promoters play an important role in the regulation of temporal and spatial expression of genes. Using high efficiency promoters to regulate the expression of specific genes of resistance genes can not only increase the ability of crops to resist stress, but also have important meanings for improving varieties. Until now, there have been few reports on the efficiency of the initiation of pGOS2 promoter of the eukaryotic translation initiation factor 1b promoter. In particular, studies on the evaluation of the initiation efficiency of the CaMV35S double-copy promoter have not been reported yet . In this study, PCR and bioinformatics techniques were used to isolate and analyze the characteristics of GOS2 promoter in rice and construct pGOS2 :: GUS binary expression vector. Agrobacterium-mediated transformation of rice embryogenic callus and transgenic rice Plants and analyzed by PCR and GUS histochemical analysis to assess the efficiency of p GOS2 in transgenic rice leaf tissue. Bioinformatics analysis showed that the cloned GOS2 promoter has a total length of 3 155 bp and possesses some typical eukaryotic cis-elements such as TATA-box, AGA-motif, CAAT-box and GCGC-repeat The GOS2 promoter core sequence was located at -43 bp to +4 bp with a score of 0.97. The results of GUS histochemical analysis showed that GUS signal was not found in the negative control group, while GUS activity in the transgenic group showed different degrees of GUS activity, and the priming efficiency of pGOS2 was much higher than that of the single and double copy CaMV35S promoter. The results of this study confirmed that the activity of GOS2 promoter in rice leaf tissue was much higher than that of the enhanced CaMV35S promoter and provided a feasible method and a new method for the development and screening of rice molecular breeding and related promoters in the future Vision.