红花转录物组中筛选的Unigene100791基因克隆、生物信息学分析及功能预测
Cloning and bioinformatic analyzing of Unigene100791 base on safflower transcriptome sequence
-
摘要: 目的:克隆红花Unigene100791基因,研究其表达情况;并对该基因的特性进行生物信息学分析,为后续红花代谢调控及提高红花油产量,使其满足食品工业市场需求奠定理论基础。方法:从红花转录物组数据库中筛选得到转录因子Dof相关基因Unigene100791的中间片段;利用RNAiso plus试剂法提取红花种子总RNA,利用RT-PCR方法反转录得到红花种子c DNA;采用3’Race方法从红花种子中克隆Unigene100791基因的全长序列;通过生物信息学方法对其全长基因进行分析,构建该基因与相关物种的系统进化树,研究其蛋白序列的同源性及其理化性质与结构特征,预测其生物学功能。结果:成功克隆了红花Unigene100791基因,其基因全长1274 bp;开放阅读框981 bp,编码326个氨基酸,理论分子质量约为35026.5 u,理论等电点9.5,带正电残基27个,带负电残基18个,序列含有典型的加尾信号poly(A);红花Unigene100791基因编码产物与烟草Dof2.4-1亲缘关系较近,属于Dof超家族,此蛋白存在信号肽,不存在跨膜结构。结论:成功克隆了红花Unigene100791基因,并对其编码的蛋白质进行了生物信息学分析。Abstract: Objective: To clone the Unigene 100791,study its expression,its characteristics was investigated using bioinformatics methods in order to laid a theoretical basis for subsequent metabolidm and improve the yield of safflower oil,which can meet the market demand of food industry. Methods: The fragment of Unigene 100791,related Dof transcription factors,was obtained by screening from safflower transcriptome data. Total RNA was extracted by using RNAiso plus reagent,using RT-PCR method to obtain the c DNA in safflower seeds. The full length sequence of Unigene 100791 was obtained with 3'Race method.The full length sequence of Unigene 100791 was analyzed using the bioinformatics methods,and the phylogenetic treewas constructed to study the homology,physicochemical property and structure characteristics in order to predict its biological function.Results: The 1274 bp Unigene100791 sequence in safflower was successfully cloned,which had a 981 bp ORF,encoding 326 amino acids.Its molecular weight was 35026.5 u,and the theoretical isoelectric point was 9.5,positively charged residues of 27,negatively charged residues of 18. The sequence contained a typical tailing signal poly( A). The encoding product of Unigene100791 in safflower had a closer genetic relationship with tobacco Dof2.4-1,belonging to the superfamily Dof,and this protein existed a signal peptide but had no transmembrane structure. Conclusion: The Unigene100791 was successfully cloned in safflower and used the bioinformatics approaches to analyze the encoded protein for Unigene100791.