【摘要】 HY5(ELONGATED HYPOCOTYL 5)转录因子在花青素的生物合成方面具有重要的作用。该研究以‘全红’杨叶片为材料，克隆了PdHY5基因，对其进行生物信息学分析；并通过农杆菌介导的叶盘法转化烟草，对其进行功能分析。结果显示：(1)PdHY5基因开放阅读框(ORF)长度为510 bp,共编码169个氨基酸。多序列比对和系统进化分析表明，‘全红’杨PdHY5蛋白具有bZIP家族蛋白的保守结构域，与毛果杨PtHY5蛋白亲缘关系最近。(2)成功构建过表达载体pNP1302-35S-PdHY5,经潮霉素筛选获得了4个转基因烟草株系(S1～S4)。(3)qRT-PCR结果表明，PdHY5基因在4个过表达株系中的表达量显著高于野生型(WT),且S4株系的表达量最高，S1最低；同时过表达株系的花青素生物合成途径关键基因CHS、F3H及FLS的表达水平较WT均显著上调。(4)叶片花色素苷的相对含量在转基因烟草株系S2、S3、S4中较WT显著上调，分别增加了128.23%、97.36%和134.20%。研究表明，过表达PdHY5基因调控了烟草本身花青素生物合成途径中结构基因的表达，从而促进了花青素的积累。更多还原

【Abstract】 Transcription factor HY5(ELONGATED HYPOCOTYL5) played an important role in anthocyanin biosynthesis. In this study, we firstly cloned PdHY5 gene from the Populus deltoides ‘Quanhong’ and analyzed it by bioinformatics methods, we then transformed PdHY5 gene into tobacco by Agrobacterium-mediated leaf disc method for functional analysis. The results showed that:(1) the open reading frame(ORF) of PdHY5 was 510 bp and encoded 169 amino acids. Comparison of amino acid sequences and phylogenetic analysis indicated that PdHY5 contained a conserved domain of bZIP family and had a closest relationship with PtHY5.(2) An over-expression vector pNP1302-35S-PdHY5 was successfully constructed, and 4 tobacco transgenic lines(S1-S4) were obtained by hygromycin screening.(3) qRT-PCR results showed that the expression of PdHY5 gene in the 4 overexpression lines was significantly higher than that of WT(wild type), with the highest expression of S4 and the least of S1. What’s more, the expression levels of several key genes participated in anthocyanin biosynthesis, such as CHS, F3H, and FLS in overexpression lines were significantly up-regulated compared with WT.(4) The relative content of anthocyanin was significantly higher in transgenic tobacco lines S2, S3 and S4 compared with WT, increasing by 128.23%, 97.36% and 134.20%, respectively. This study indicated that overexpression of PdHY5 gene regulated the expression of structural genes in the anthocyanin biosynthesis pathway of tobacco itself, thereby promoting the accumulation of anthocyanins.更多还原