凡纳滨对虾WAP基因重组表达及抗病功能分析  

Recombination Expression and Analysis of Disease Resistance of WAP Gene in Litopenaeus vannamei

作　　者：马春霞[1] 黄婷 卢敏 陆专灵 谢宗升 雷爱莹 陈福艳 熊建华 黎铭 MA Chun-xia;HUANG Ting;LU Min;LU Zhuan-ling;XIE Zong-sheng;LEI AI-ying;CHEN Fu-yan;XIONG Jian-hua;LI Ming(Guangxi Veterinary Research Institute/Cuangxi Key Laboratory of Veterinary Biotechnology,Guangxi Nanning 530001,China;Guangxi Academy of Fisheries Sciences,Guangxi Nanning 530021,China/Guangxi Key Laboratory of Aquatic Genetic Breeding and Healthy Aquaculture;Guangxi Key Laboratory of Beibu Gulf Marine Biodiversity Conservation/School of Ocean Beibu Gulf University,Guangxi Qinzhou 535011,China)

机构地区：[1]广西兽医研究所/广西兽医生物技术重点实验室,广西南宁530001 [2]广西水产科学研究院/广西遗传育种与健康养殖遗传重点试验室,广西南宁530021 [3]广西北部湾海洋生物多样性养护重点实验室/北部湾大学海洋学院,广西钦州535011

出　　处：《西南农业学报》2020年第10期2398-2404,共7页Southwest China Journal of Agricultural Sciences

基　　金：广西重点研发计划项目(桂科AB18221115);国家现代农业产业技术体系建设项目广西对虾产业创新团队专项(nycytxgxcxtd-14-05);广西科技重大专项(科AA17204095-1)。

摘　　要：【目的】针对凡纳滨对虾白便病的危害现状,克隆其WAP基因并分析其组织表达特性及抗病功能,为凡纳滨对虾抗病药物开发提供参考依据。【方法】设计WAP基因特异性引物,用RT-PCR检测WAP基因在凡纳滨对虾血细胞、眼柄、鳃、肝胰腺、心脏、胃、肌肉、神经、肠、后盲囊和表皮等11种组织的表达分布情况;采用实时荧光定量PCR检测患白便病凡纳滨对虾鳃和肠道组织的WAP基因表达量;重组表达WAP蛋白,使用ELISA分析其与细菌多糖LPS(脂多糖)、PGN(肽聚糖)和LTA(脂磷壁酸)的结合能力。【结果】WAP基因在11个凡纳滨对虾组织均有表达,其中,以在血细胞中的表达量最高,在表皮、眼柄和肝胰腺的表达量表达较低;WAP基因在感染白便病凡纳滨对虾鳃和肠道组织的表达量显著高于健康凡纳滨对虾(P<0.05),分别为健康凡纳滨对虾的5.9258和1.5147倍;WAP融合蛋白与细菌多糖LPS、PGN和LTA具有直接结合作用,细菌多糖的结合能力表现为PGN>LPS>LTA。【结论】WAP基因与凡纳滨对虾抗病免疫功能密切相关,WAP融合蛋白在开发凡纳滨对虾抗白便病药物方面具有潜在应用价值。【Objective】Because of the serious disease problem of Litopenaeus vannamei,it is important to study the disease resistance genes of L.vannamei.In this study,the WAP gene of L.vannamei was cloned and its tissue expression and anti-disease function were analyzed to provide reference for the development of anti-disease drugs for L.vannamei.【Method】The specific primers of WAP gene were designed,and the expression and distribution of the WAP gene in 11 tissues including blood cells,eye stem,gills,hepatopancreas,heart,stomach,muscle,nerve,intestine,cecum and epidermis were detected by RT-PCR.The WAP expression in gills and intestinal tissues of L.vannamei was detected by fluorescence quantitative PCR.The recombinant WAP protein was produced and the binding ability of recombinant WAP to LPS,PGN and LTA was analyzed by ELISA.【Result】It was found that WAP gene was expressed in all 11 tissues tested,with the highest expression in blood cell and the lowest expression in epidermis,eye stalk and hepatopancreas.The WAP gene expression was significantly increased in gill and intestinal tissues of infected L.vannamei compared with that of normal L.vannamei(P<0.05).The expression of WAP gene in the gill and intestinal tissues of infected L.vannamei were as 5.9258 and 1.5147 times as that of healthy L.vannamei,respectively.The recombinant WAP fusion protein could bind to bacterial polysaccharides LPS,PGN and LTA directly,and the order of binding ability was PGN>LPS>LTA.【Conclusion】The WAP gene was closely related to the disease resistance and immune function of L.vannamei,WAP fusion protein has potential in developing anti white feces drugs for L.vannamei.

关 键 词：凡纳滨对虾 WAP基因 白便病 抗病功能 

分 类 号：S945.1[农业科学—水产养殖]