褐牙鲆白细胞介素21基因克隆及表达分析  

Cloning and Expression of Interleukin 21 Gene in Bastard Halibut Paralichthys olivaceus

作　　者：薛婷婷 郑津辉 周密 潘宝平 孙金生 高虹 XUE Tingting;ZHENG Jinhui;ZHOU Mi;PAN Baoping;SUN Jinsheng;GAO Hong(Tianjin Key Laboratory of Animal and Plant Resistance,College of Life Sciences,Tianjin Normal University,Tianjin 300387,China)

机构地区：[1]天津师范大学生命科学学院天津市动植物抗性重点实验室

出　　处：《水产科学》2020年第2期151-159,共9页Fisheries Science

基　　金：天津市自然科学基金重点项目(14JCZDJC34200,19JCZDJC34300)

摘　　要：利用转录组测序数据信息和PCR技术克隆褐牙鲆白细胞介素21(IL-21)基因的编码区序列,并进行生物信息学分析及表达模式分析。试验结果显示,白细胞介素21基因开放阅读框429 bp,编码142个氨基酸,分子量16.0 ku,在N端具有19 bp信号肽序列。分子系统发育树分析表明,褐牙鲆白细胞介素21蛋白与尖吻鲈以及深裂眶锯雀鲷白细胞介素21蛋白亲缘关系最近。实时荧光PCR结果显示,白细胞介素21基因在所检测的褐牙鲆7种健康组织中均有表达,其中肝脏的表达量最高。用提取的迟钝爱德华氏菌脂多糖免疫刺激褐牙鲆后,白细胞介素21基因的表达在头肾和脾组织中产生不同程度上调,表明该基因参与了脂多糖诱导的免疫应答。利用RNA干扰技术沉默髓样分化因子(MyD88)基因后,白细胞介素21基因的表达出现下调,当再用提取的迟钝爱德华氏菌脂多糖免疫刺激后,MyD88和白细胞介素21基因的表达又均开始逐渐回升,暗示白细胞介素21与MyD88通路具有一定关联性。The coding sequence of interleukin-21(IL-21) gene was cloned and expressed in bastard halibut Paralichthys olivaceus by using transcriptome sequencing data and PCR technology, and analyzed by bioinformatics analysis. The results showed that the open reading frame of IL-21 gene was 429 bp, encoding 142 amino acids with a molecular weight of 16.0 ku, and had a 19 bp signal peptide sequence at the N-terminal. Molecular phylogenetic tree indicated that IL-21 protein of bastard halibut had the closest relationship with IL-21 of perch Lates calcarifer and Stegastes partitus. Real-time fluorescent PCR revealed that IL-21 gene was expressed in all seven healthy tissues of bastard halibut, with the maximal expression level in liver. The expression of IL-21 was found to be up-regulated in head kidney and spleen of bastard halibut immunized with the extracted lipopolysaccharide from Edwardsiella tarda, indicating that IL-21 was involved in the immune response induced by lipopolysaccharide. When MyD88 gene was silenced by RNA interference, the expression of IL-21 was down-regulated, and the expression of MyD88 and IL-21 genes began to increase gradually in the bastard halibut immunized with the extracted lipopolysaccharide from E. tarda, indicating that that IL-21 is related to MyD88 pathway.

关 键 词：白细胞介素21 褐牙鲆 迟钝爱德华氏菌 基因表达 荧光实时定量PCR 

分 类 号：S965.199[农业科学—水产养殖]