杂交石斑鱼干扰素调节因子3(IRF3)基因的克隆及表达分析  

Cloning and Expression Analysis of Interferon Regulatory Factor 3(IRF3) Gene in Hybrid Grouper

作　　者：卢晓颖 黄宝松 马骞 陈刚[1] 王忠良[1] 黄建盛[1] ERIC AMENYOGBE 谢瑞涛[1,2] 邓文鑫 LU Xiao-ying;HUANG Bao-song;MA Qian;CHEN Gang;WANG Zhong-liang;HUANG Jian-sheng;ERIC AMENYOGBE;XIE Rui-tao;DENG Wen-xin(College of Fishery,Zhanjiang 524088;Guangdong Evergreen Feed Industry Co.Ltd,Zhanjiang 524000;Guangdong Yuehai Feed Group Co.Ltd,Zhanjiang 524088)

机构地区：[1]广东海洋大学水产学院,湛江524088 [2]广东恒兴饲料实业股份有限公司,湛江524000 [3]广东粤海饲料集团有限公司,湛江524088

出　　处：《生物技术通报》2019年第10期144-151,共8页Biotechnology Bulletin

基　　金：2016广东省省级科技计划项目(2016B020201009)

摘　　要：干扰素调节因子家族(IRFs)具有抗病毒、免疫调节功能,干扰素调节因子3(Interferon regulatory factor 3,IRF-3)是IRFs家族中的一员,也是免疫相关因子。为了解IRF3基因在杂交石斑鱼(Epinephelus.fuscoguttatus,♀)×清水石斑鱼(E.polyphekadion,♂)应对外源病毒刺激的免疫反应,利用cDNA末端快速扩充(Rapid-amplification of cDNA ends,RACE)技术克隆了杂交石斑鱼IRF3基因,该基因cDNA全长为2 529 bp,包含5'非编码区(5'-UTR)325 bp,3'非编码区(3'-UTR)916 bp,开放阅读框(Open reading frame,ORF)1 377 bp,可编码458个氨基酸。氨基酸序列包含N-末端DNA结合区(N-terminal DNA binding region,DBD)(1-108 aa)、1个C-末端干扰素相关区(C-terminal interferon related region,IAD)(255-435 aa)及色氨酸富含区(Tryptophan rich region,SRD)(440-450 aa)3个结构域。系统进化分析结果显示,杂交石斑鱼IRF3与花鲈(Lateolabrax maculatus)IRF3聚为一支,亲缘关系较近。利用实时荧光定量PCR检测IRF3基因在杂交石斑鱼肝、胃、鳃、肠和脾脏等9种组织表达情况,以及在外周血淋巴细胞(Peripheral blood lymphocytes,PBL)的时序性表达情况,结果显示,IRF3基因在9种组织中均有表达,肝、胃和肠中的表达量明显高于其他组织,头肾表达量最低。PBL在PolyI:C刺激1 h后IRF3基因表达量逐渐升高,4 h时表达量达到最大值(约为对照组的9.3倍),8 h后逐渐下降。Interferon regulatory factor(IRF)family has antiviral and immunoregulatory functions.Interferon regulatory factor 3(IRF-3)is a member of the IRF family and is also an immune-related factor.To understand the immune response of gene IRF3 in the hybrid grouper(Epinephelus fuscoguttatus,♀)×(E.polyphekadion,♂)to exogenous virus stimulation,the IRF3 gene of the hybrid grouper was cloned by RACE(rapid-amplification of cDNA ends).The full-length cDNA of this gene is 2 529 bp,including 5'non-coding region(5'-UTR)325 bp,3'non-coding region(3'-UTR)916 bp,open reading frame(ORF)1 377 bp,and encoding 458 amino acids.The amino acid sequence comprises of 3 domains,i.e,one N-terminal DNA binding region(DBD)(1-108 aa),one C-terminal interferon related region(IAD)(255-435 aa),and a tryptophan rich region(SRD)(440-450 aa).Phylogenetic analysis showed that the hybrid grouper’s IRF3 and the Japanese sea perch’s(Lateolabrax maculatus)IRF3 clustered into one,thus the genetic relationship was close.Real-time quantitative PCR was used to detect the expression of gene IRF3 in 9 tissues of liver,stomach,gill,intestine,spleen of hybrid grouper,etc.as well as the temporal expression of peripheral blood lymphocytes(PBL).The results showed that gene IRF3 expressed in 9 tissues,and the expression in the liver,stomach and intestine was significantly higher than that in other tissues,and the expression in head and kidney was the lowest.The expression of gene IRF3 in PBL increased gradually after stimulation for 1 h at PolyI:C,and reached the maximum at 4 h(about 9.3 times of the control group),and gradually decreased after 8 h.

关 键 词：杂交石斑鱼 干扰素调节因子3 基因克隆 外周血淋巴细胞 PolyI:C 

分 类 号：R73[医药卫生—肿瘤]