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AP2/ERF转录因子家族广泛存在于植物中,参与植物细胞周期、生长发育以及生物和非生物胁迫相关基因表达调控。本文利用油菜UniGene数据库,以拟南芥AP2/ERF-B3亚族转录因子保守序列为信息探针,分离得到2个油菜AP2/ERF-B3亚族的转录因子BnaERFB3-1和BnaERFB3-2。通过PCR和RT-PCR方法分别从双低甘蓝型油菜沪油15的DNA和cDNA中克隆了上述基因。序列分析显示,克隆的BnaERFB3-1-Hy15和BnaERFB3-2-Hy15转录因子与电子克隆的基因序列差异很小,均只有1个氨基酸位点不同,且都没有内含子。从氨基酸序列的相似性、组成成分、理化性质、疏水性/亲水性、序列比对、进化树、功能域、二级结构、三级结构、无序化特性等方面进行了预测和较为全面的分析。结果显示BnaERFB3-1-Hy15和BnaERFB3-2-Hy15是亲水性蛋白,在蛋白质的三级结构上与AtERF5相似,BnaERFB3-1-Hy15和BnaERFB3-2-Hy15蛋白无序化程度大于拟南芥AtERF5。通过分析EST丰度显示,BnaERFB3-1的表达集中在种子中,而BnaERFB3-2的表达则集中在根中。另外,将上述基因分别构建入酵母表达载体和植物双元表达载体,为深入研究该基因在油菜抗逆调控中的作用奠定了基础。
The AP2 / ERF family of transcription factors is widespread in plants and is involved in the regulation of cell cycle, growth and development and the expression of genes involved in biotic and abiotic stress. In this study, we used the UniGene database of rapeseed to study the transcriptional factors BnaERFB3-1 and BnaERFB3-2 of AP2 / ERF-B3 subfamily from two Arabidopsis AP2 / ERF-B3 subfamily transcription factor conserved sequences as informative probes. The above genes were cloned by PCR and RT-PCR respectively from DNA and cDNA of double-low Brassica napus Huyou 15. Sequence analysis showed that the cloned BnaERFB3-1-Hy15 and BnaERFB3-2-Hy15 transcription factors and the electronic cloning gene sequence difference is very small, only one amino acid sequence difference, and no introns. From the amino acid sequence similarity, composition, physical and chemical properties, hydrophobicity / hydrophilicity, sequence alignment, evolutionary tree, functional domain, secondary structure, tertiary structure, disorder characteristics and other aspects of the prediction and more comprehensive Analysis. The results showed that BnaERFB3-1-Hy15 and BnaERFB3-2-Hy15 are hydrophilic proteins, which are similar to AtERF5 in the tertiary structure of proteins. The degree of disorder of BnaERFB3-1-Hy15 and BnaERFB3-2-Hy15 proteins is greater than that of Arabidopsis thaliana AtERF5. By analyzing EST abundance, the expression of BnaERFB3-1 was concentrated in seeds, while the expression of BnaERFB3-2 was concentrated in the roots. In addition, the above genes were constructed into yeast expression vector and plant binary expression vector, which laid the foundation for further study on the role of this gene in the resistance regulation of rapeseed.