烟草NtMYB102转录因子基因克隆及生物信息学分析

Gene cloning and bioinformatics analysis of NtMYB102 transcription factor in Nicotiana tabacum

为解析NtMYB102转录因子的结构并预测其功能,采用同源克隆和Sanger测序法,从烟草品种K326中获得NtMYB102基因的CDS序列,基于该序列分析NtMYB102基因编码蛋白的理化性质、结构域和基因上游顺式作用元件。从NCBI SRA数据库中下载烟草接种青枯雷尔氏菌(Ralstonia solanacearum)后的转录组原始数据,分析NtMYB102基因的表达量变化,并采用实时荧光定量PCR法分析烟草接种蚜虫后NtMYB102基因的相对表达量变化。结果表明:①NtMYB102基因全长1131 bp,编码的NtMYB102蛋白由376个氨基酸组成,具有典型的MYB特征结构域,是一个无信号肽、无跨膜结构、具有磷酸化位点的亲水蛋白。②NtMYB102基因上游2000 bp有多个与逆境胁迫、病原响应相关的调控元件。③抗烟草青枯病品种岩烟97接种青枯雷尔氏菌后36 h,NtMYB102基因表达量显著低于0 h,表明该基因可能在烟草响应青枯雷尔氏菌的侵染中起调控作用。④接种蚜虫后,烟草品种中烟100 NtMYB102基因的相对表达量显著提高,表明该基因可能参与烟草对蚜虫取食的响应。

To characterize the structure and function of NtMYB102 tobacco transcription factor,the CDS of NtMYB102 gene was cloned from tobacco cultivar K326 by homology-based cloning and Sanger sequencing.Physicochemical characteristics,domains of NtMYB102 protein and upstream sequence of NtMYB102 gene were analyzed by online tools.The expression level of NtMYB102 gene after Ralstonia solanacearum infection was analyzed based on transcriptom raw data downloaded from NCBI SRA.Relative expression level of NtMYB102 gene after being infested by aphids was analyzed by qRT-PCR.The results showed that:1)The NtMYB102 gene was 1131 bp in length and encoded a protein consisting of 376 amino acids.The NtMYB102 protein had typical MYB domains,and was a hydrophilic protein with phosphorylation sites while lacking signal peptides and transmembrane structure.2)The 2000 bp sequence upstream of the NtMYB102 gene contained several regulatory elements associated with stress and pathogen response.3)At 36 h after infection by R.solanacearum,the relative expression level of the NtMYB102 gene of Yanyan 97(resistant to tobacco bacterial wilt)was significantly lower than that at 0 h,suggesting that this gene may be involved in coping with infection by R.solanacearum.4)The relative expression level of the NtMYB102 gene of Zhongyan 100 was significantly increased after aphid infestation,indicating that this gene may be involved in the response of tobacco to aphids.