农业生物技术学报2017，Vol.25Issue(6)：861-873,13.DOI:10.3969/j.issn.1674-7968.2017.06.001
尼罗罗非鱼TRIM16和TRIM25基因的克隆及表达分析
Cloning and Expression Analysis of TRIM16 and TRIM25 Genes from Nile Tilapia (Oreochromis niloticus)
摘要
Abstract
The ubiquitin ligase E3 family plays an important role in the innate immunity of vertebrate,and many members of tripartite motif (TRIM) family have fimction as an E3 ligase in the ubiquitin modification based on the existence of the conserved RING finger domain.In order to evaluate the immune function of TRIM16 and TRIM25 genes from Nile tilapia (Oreochromis niloticus),the full-length cDNA sequences of TRIM16 and TRIM25 genes from were obtained by reverse transcription PCR and RACE.In addition,the gene structure and protein secondary structure were also analyzed.qRT-PCR was used for analyzing the expression of TRIM16 and TRIM25 genes in tissues and the process of embryonic development,and the response to Streptococcus agalactiae infection.Results showed that the TRIM16 gene (GenBank accession No.KY746714)was 2 314 bp in total length which contained an ORF of 1 677 bp encoding 558 amino acid residues.The 5'-UTR and 3'-UTR were 13 and 624 bp,respectively.TRIM25 gene (GenBank accession:No.KY968697)was 2 748 bp in total length which contained an ORF of 1 677 bp encoding 558 amino acid residues.The 5'-UTR and 3'-UTR were 21 and 1 050 bp,respectively.TRIM16 and TRIM25 genes were all without introns.The results of putative protein indicated that the TRIM16 and TRIM25 had conserved structures of the TRIM family,such as the RING finger domain and the B-box domain.The putative protein of TRIM16 had 29.0％～ 92.6％ identities with other teleosts and mammals.The putative protein of TRIM25 had 20.0％～88.1％ identities with other teleosts and mammals.The putative protein of TRIM16 and TRIM25 exhibited the highest identity with Maylandia zebra (92.6％ and 88.1％,respectively).Tissue distribution analysis revealed that TRIM16 and TRIM25 genes expressed in all tested tissues with the highest expression level in blood and the lowest expression level in liver.The expression levels of TRIM16 and TRIM25 in the blood were 60.46 and 274.07 folds compared with that in the liver,respectively.Both TRIM16 and TRIM25 genes expressed during the embryonic development of Nile tilapia,which suggested that they played an important role in the immune process of Nile tilapia in the early stage of embryonic development.Upon stimulation with intraperitoneal injection with Streptococcus agalactiae,the expression level of TRIM16 and TRIM25 genes were up-regulated in all test tissues (intestine,spleen,gill,kidney and blood) at the same times.The highest expression levels of TRIM16 gene in intestinal,spleen,gill,kidney and blood were 1.30,2.09,1.61,7.81 and 6.05 folds compared with 0 h (control group),respectively.The highest expression levels of TRIM25 gene in the intestine,spleen,gill,kidney and blood were 11.13,1.22,1.26,61.41 and 77.80 folds compared with 0 h (control group),respectively.The results showed that TRIM16 and TRIM25 genes played an important role in the immunoreaction of Nile tilapia against S.agalactia infection.This study provides a theoretical basis for further understanding of the anti-infective immune mechanism of Nile tilapia and exploring new ways of disease prevention and control.关键词
尼罗罗非鱼/三重基序(TRIM)蛋白家族/基因克隆/无乳链球菌Key words
Nile tilapia/Tripartite motif (TRIM) protein family/Gene cloning/Streptococcus agalactiae分类
农业科技引用本文复制引用
郑建美,高风英,卢迈新,刘志刚,曹建萌,可小丽,王淼..尼罗罗非鱼TRIM16和TRIM25基因的克隆及表达分析[J].农业生物技术学报,2017,25(6):861-873,13.基金项目
现代农业产业技术体系专项(CARS-49)、广东省创新载体建设项目(No.2013B090700004)和广州市科技计划项目产学研协同创新重大专项(No.201508020046) （CARS-49）
