樊云霞,简纪常,鲁义善,吴灶和

• 1:广东海洋大学水产学院
• 2:广东省水产经济动物病原生物学及流行病学重点实验室
• 3:广东省教育厅水产经济动物病害控制重点实验室
• 4:仲恺农业工程学院

摘要(Abstract)：

根据红笛鲷(Lutjanus sanguineus)CD40基因cDNA全长序列设计2对特异性引物,利用RT-PCR技术从红笛鲷头肾组织中扩增出CD40 ORF序列和去信号肽序列,分别与原核表达载体pET-32a(+)相连,构建原核重组表达质粒,命名为pET32-CD40和pET32-△CD40,并分别转化至大肠杆菌Rosetta,经IPTG诱导后,分别于54 ku和53 ku处有清晰的目的蛋白条带。融合蛋白的表达效率最佳的表达条件,pET32-CD40是诱导温度37℃,起始D(600nm)为0.4,IPTG浓度为0.08 mmol/L,诱导5 h;pET32-△CD40是诱导温度37℃,起始D(600nm)为0.6,IPTG浓度为0.10 mmol/L,诱导6 h。在各自优化的表达条件下,pET32-△CD40融合蛋白表达量较pET32-CD40融合蛋白高。RNAstructure软件分析发现,CD40的mRNA 5′端序列形成复杂且较为稳定的二级结构,不利于翻译的起始,表明信号肽序列的存在可能对CD40基因在原核细胞中的表达有一定的影响。

关键词(KeyWords)： 红笛鲷;CD40基因;原核表达

Abstract:

Keywords:

基金项目(Foundation): 国家自然科学基金项目(41240041);; 广东省科技厅国际合作项目(2012B050600029)

作者(Author): 樊云霞,简纪常,鲁义善,吴灶和

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