甘蓝型油菜中一类AP2／ERF转录因子的克隆和生物信息学分析

In Silico Cloning and Bioinformatic Analyzing of AP2/ERF Transcription Factor Genes from Brassica napus L.

AP2/ERF转录因子家族是植物中广泛存在的一类转录因子,AP2/ERF这类转录因子主要参与植物的细胞周期、生长发育以及生物和非生物胁迫相关基因的表达调控。由于拟南芥和油菜同属于芸薹属,具有相似的基因信息,利用油菜UniGene数据库,以拟南芥ERF转录因子保守序列为探针,通过电子克隆从一个UniGene C luster中分离得到三个同类的AP2/ERF转录因子,从cDNA序列、氨基酸序列的相似性、组成成分、理化性质、疏水性/亲水性分析、序列比对、进化树、功能域、二级结构、三级结构、无序化特性进行了预测和较为全面的分析。结果显示油菜来源的BnaERF1、BnaERF2和BnaERF3属于AP2/ERF转录因子的B-2亚族,是亲水性蛋白,在蛋白质的三级结构上与AtERF1相似。蛋白质无序化分析发现,油菜BnaERF1、BnaERF2和BnaERF3无序化程度小于拟南芥AtERF1。设计引物通过PCR和RT-PCR方法分别从甘蓝型双低油菜沪油15幼苗的DNA和cDNA中扩增了上述基因,初步分析BnaERF2没有内含子,BnaERF1和BnaERF3有内含子。另外,通过EST丰度分析显示,该类转录因子的表达最高峰在种子中,其次为花,在芽、茎和分生组织中没有检测出表达的存在。

Plants are constantly challenged by various biotic and abiotic stresses, such as disease, drought, high salinity, and low temperature in their natural environment, which cause adverse effects on the growth of plants and serious reductions in the quantity and quality of crops. AR2/ERF is a large family of transcription factors. Genes in the ERF family encode transcriptional regulators with a variety of functions involved in the developmental and physiological processes in plants. Three AP2/ERF family transcriptional regulators（ BnaERF1, BnaERF2 and BnaERF3 ）were isolated from Brassica napus by in silico cloning method using the conserved domain amino acid sequence of Arabidopsis thaliana ERF as probe. Then, cDNA and deduced amino acid sequence, composition, hydrophobicty and hydrophilicity, physical and chemical characterization, phylogenetic tree, conserved domain sequences, function domain, molecular modeling, and folding state were predicted and analyzed. Then BnaERF1, BnaERF2 and BnaERF3 genes from B. napus L. Huyoul5 were isolated by RT-PCR and PCR using cDNA and DNA as template.