尼罗罗非鱼淋巴毒素α基因克隆与表达分析Cloning and Expression Analysis of Gene Lymphotoxin α in Nile Tilapia(Oreochromis Niloticus)
邹子鸿,郑琦,黄瑜,汤菊芬,王蓓,鲁义善,简纪常,蔡佳
摘要(Abstract):
【目的】克隆尼罗罗非鱼(Oreochromis niloticus)淋巴毒素α基因(lymphotoxin alpha, LTα),分析该基因的组织分布,探讨该基因在抗菌抗病毒免疫过程中的重要作用。【方法】通过RACE技术获得尼罗罗非鱼淋巴毒素α基因(On-LTα)的cDNA全长,通过生物信息学分析其序列结构特征;利用实时荧光定量PCR检测基因的组织分布特征,以及健康尼罗罗非鱼感染无乳链球菌后该基因的表达变化,并进一步分离出尼罗罗非鱼非特异性毒性细胞(NCC),检测脂多糖(LPS)和聚肌胞苷酸(Poly I:C)刺激后,On-LTα在NCC中的表达变化。【结果】On-LTα基因序列全长为2699 bp,包含705 bp开放阅读框(ORF),编码234个氨基酸(GenBank登录号:MK770358)。该蛋白属于跨膜蛋白,具有肿瘤坏死因子(TNF)家族保守结构域。实时荧光定量结果表明,On-LTα在健康尼罗罗非鱼11种组织中均有表达,在脾脏中的表达量最高;无乳链球菌刺激后,该基因在脾脏中的表达量显著升高;LPS与PolyI:C刺激后,On-LTα在NCC中表达量也显著上调。【结论】On-LTα参与了尼罗罗非鱼抗菌抗病毒的免疫应答过程。
关键词(KeyWords): 尼罗罗非鱼;淋巴毒素α基因;基因克隆;基因表达;非特异性细胞毒性细胞(NCC)
基金项目(Foundation): 国家自然科学基金(31302226,31572651);; 广东海洋大学优秀硕士学位论文培育项目(201833)
作者(Author): 邹子鸿,郑琦,黄瑜,汤菊芬,王蓓,鲁义善,简纪常,蔡佳
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