作　　者： ; ; ; ; ; ; ; ;

机构地区： 广西水产研究所

出　　处： 《水生生物学报》 2010年第5期949-955,共7页

摘　　要： 颗粒蛋白前体(Progranulin,PGRN)在先天免疫反应调控及个体生长发育过程中均有重要作用。通过对罗非鱼外周血白细胞全长cDNA文库筛选得到的序列进行生物信息学分析,获得罗非鱼Pgrn全长cDNA序列(GenBank登录号为GQ241348)。该cDNA克隆总长843bp,包含一个完整的开放阅读框,编码206个氨基酸,其中推定信号肽20个氨基酸,两个GRN重复单位均56个氨基酸。研究采用实时定量PCR(Real-time RT-PCR)方法对感染海豚链球菌后奥尼罗非鱼、尼罗罗非鱼、奥利亚罗非鱼和吉富罗非鱼4种组织(脑、肝脏、脾脏和头肾)Pgrn mRNA表达情况进行分析。结果显示,Pgrn mRNA在攻毒后4种罗非鱼4种组织中表达均有上调趋势,并且在脾中的表达量最高,提示PGRN在鱼类先天免疫反应调控中起重要作用。另外,奥尼罗非鱼在感染海豚链球菌后6h的脑和肝脏、6h和12h的脾脏和头肾中Pgrn mRNA表达均下调,然后表达升高,这种表达变化在其他三种鱼中不明显,这也许是奥尼罗非鱼抗病力较强的一个原因。研究为从分子水平探讨PGRN在罗非鱼先天免疫反应中的作用机制提供了数据,也为罗非鱼的抗病选育提供了参考分子标记。 Progranulin （PGRN） is an epithelial tissue growth factor （also known as proepithelin, acrogranin and PC-cell-derived growth factor） which plays an important role in innate immunity regulation and growth development. The full-length cDNA of Pgrn （GenBank Accession number： GQ241348） was obtained by screening the full-length cDNA library of the peripheral blood leucocytes from Oreochromis niloticus. The full size of the cDNA was 843 bp, containing one complete open reading frame encoding a peptide of 206 amino acid residues （aa） that contains a presumptive signal peptide （20 aa） and two repeat units of granulin （GRN, both 56 aa）. The two predicted GRN peptides were arranged in tandem repeats interrupted by a GAP peptide. The deduced amino acid sequences of the two predicted GRN peptides were consistent with the motif of X2-3CX5-6CX5CCX8CCX6CCX5CCX4CX5-6CX2. The positions of the cysteine residues were highly conserved in the GRN consensus sequence of tilapia, carp, zebra fish and human. respectively. GRN1 and GRN2 of O. niloticus and O. mossambicus had two and one amino acid variation sites respectively. Furthermore, phylogenetic analysis of GRN1 and GRN2 from tilapia, carp, zebra fish and human revealed that while tilapia GRN2 was closer to carp and zebra fish GRNs, tilapia GRN1 was the closest to human GRN2 and GRN3. In the present study, four species of tilapia （O. niloticus×O. aureus, O. niloticus, O. aureus and GIFT） were divided into infection and control groups with 50 tilapias in each group. The infection dose for each infection group （2.0×107 CFU） was the median lethal dose （LD50） for O. niloticus×O. aureus based on our previous results. The tilapias in the control group were intraperitoneally injected with equal volume of normal saline. Occurrence of disease and mortality was observed and recorded. Mortality in all the tilapia species was observed within 25 days of infection; more specifically, the mortality appeared to reached peak during day 4-6, and completely cea

关 键 词： 罗非鱼 实时定量 抗病育种 基因表达分析

领　　域： [生物学]