马氏珠母贝Nrf2基因的克隆及其在不同免疫刺激和营养条件下的表达变化  

Cloning and expression analysis of Nrf2 gene in pearl oyster Pinctada fucata martensii exposed to different immune stimulation and nutritional conditions

作　　者：蔡彩霞 廖永山 杨创业[1,2,3] 郑哲 邓岳文[1,2,3] 王庆恒 CAI Caixia;LIAO Yongshan;YANG Chuangye;ZHENG Zhe;DENG Yuewen;WANG Qingheng(College of Fisheries, Guangdong Ocean University, Zhanjiang 524088, China;Pearl Breeding and Processing Engineering Technology Research Centre of Guangdong Province, Zhanjiang 524088,China;Guangdong Provincial Engineering Laboratory for Mariculture Organism Breeding, Zhanjiang 524088, China)

机构地区：[1]广东海洋大学水产学院,广东湛江524088 [2]广东省珍珠养殖与加工工程技术研究中心,广东湛江524088 [3]广东省海水养殖生物育种工程实验室,广东湛江524088

出　　处：《大连海洋大学学报》2022年第2期202-211,共10页Journal of Dalian Ocean University

基　　金：广东省基础与应用基础研究基金委员会青年基金项目(2019A1515111026,2020A1515010691,2021A1515011199);国家自然科学基金(32002369);广东省普通高校青年创新人才类项目(2019KQNCX043);广东省农业农村厅现代农业产业技术创新团队专项(2020KJ146);财政部和农业农村部:国家现代农业产业技术体系资助(CARS-49)。

摘　　要：为探究核因子E2相关因子2(nuclear factor erythroid 2 related factor 2,Nrf2)基因在马氏珠母贝Pinctada fucata martensii中的作用,克隆获得了马氏珠母贝Nrf2基因的完整CDS区,分析了其序列特征,并运用实时荧光定量PCR分析Nrf2在不同组织、不同免疫刺激和营养条件下的表达模式。结果表明:马氏珠母贝Nrf2基因开放阅读框为2652 bp,编码883个氨基酸,预测该蛋白的相对分子质量为100150,Nrf2蛋白靠近C端处具有一个BRLZ结构域;系统进化分析显示,马氏珠母贝Nrf2氨基酸序列与长牡蛎Crassostrea gigas等双壳贝类聚为一支,亲缘关系较近;荧光定量PCR分析显示,Nrf2基因在马氏珠母贝肠道、足、外套膜、闭壳肌、鳃、肝胰腺、性腺组织中均有表达,其中,肝胰腺、肠道和鳃中的表达量显著高于其他组织(P<0.05);贝体受到LPS、PGN、Poly I:C刺激后,Nrf2在48 h时显著上调(P<0.05),且表达量最高,分别为0 h PBS对照组的3.8、2.8、3.1倍;以螺旋藻粉、酵母粉和豆粕为主要蛋白源的饲料组中Nrf2基因表达量显著高于小球藻粉和玉米粉组(P<0.05)。研究表明,Nrf2可能参与马氏珠母贝免疫应答过程,这为分析Nrf2的生物学功能及其在机体免疫调节中的作用提供了数据资料。To explore the role of nuclear factor erythroid 2 related factor 2(Nrf2)gene in pearl oyster Pinctada fucata martensii,the complete CDS region of Nrf2 gene was cloned,sequence characteristics and the expression pattern of Nrf2 were detected in different tissues of the pearl oyster by qRT-PCR technology.Then,the expression trends of Nrf2 gene were analyzed in the pearl oysters injected with 100μL of 10μg/mL phosphate buffer solution(PBS)(control group),and 100μL of 10μg/mL lipopolysaccharide(LPS),peptidoglycan(PNG),and polyinosinic acid(Poly I:C),respectively,and fed isonitrogen and isolipid diets containing different protein sources including Chlorella sp.powder,Spirulina platensis powder,yeast powder,soybean meal and corn gluten powder.The Nrf2 gene contained an open reading frame of 2652 bp,and encoded a total of 883 amino acids with relative molecular mass of 100150.There was a BRLZ domain in the C-terminal of Nrf2 amino acids.Phylogenetic analysis revealed that Nrf2 of pearl oyster was clustered with other bivalves like Crassostrea gigas.In addition,qRT-PCR analysis showed that Nrf2 gene widely existed in tissues including intestine,foot,gonad,mantle,gill,adductor muscle,and hepatopancreas.The expression level of Nrf2 was significantly upregulated under different stimulation including LPS,PGN and Poly I:C at 48 h(P<0.05),which was 3.8,2.8 and 3.1 times higher than that in the control group at 0 h,respectively.Expression level of Nrf2 was significantly higher in the pearl oyster fed the diets containing Spirulina platensis powder,yeast powder and soybean meal diets than that diets containing Chlorella sp.powder and corn gluten powder(P<0.05).The findings indicate that Nrf2 may be involved in the immune response of the pearl oyster,and provide theoretical knowledge for the analysis of the biological functions of Nrf2 and its role in immune regulation.

关 键 词：马氏珠母贝 NRF2 免疫刺激 人工配合饲料 

分 类 号：S917.4[农业科学—水产科学]