凡纳滨对虾ARMC8基因的克隆及表达
发布时间：2019-04-25 23:26
【摘要】：为了研究含Armadillo重复蛋白8在凡纳滨对虾抗病感染过程中所起的免疫作用,本实验采用RACE-PCR技术首次克隆得到凡纳滨对虾ARMC8基因(LvARMC8,Gen Bank注册号:KX058562)的c DNA序列全长,并利用在线软件进行生物信息学分析;采用实时荧光定量PCR(Rt-PCR)技术对LvARMC8基因在凡纳滨对虾不同组织及感染白斑综合征病毒和副溶血弧菌过程中的表达变化特征进行分析。结果显示,LvARMC8基因全长为2917bp,其中开放阅读框长2046 bp,编码681个氨基酸,5′非编码区长49 bp,3′非编码区长822 bp。预测分析显示LvARMC8编码的蛋白质含有6个ARM结构域。同源性分析发现,LvARMC8基因与内华达古白蚁ARMC8基因的相似度最高,为71%。系统进化分析结果显示,LvARMC8和多种无脊椎动物ARMC8聚为一支,其中与昆虫类动物赤拟谷盗、致倦库蚊和柑橘凤蝶的亲缘关系最近。Rt-PCR分析发现,LvARMC8基因在凡纳滨对虾多个组织中均能检测出,在表皮中表达量最高,眼柄中表达量最低。WSSV感染后12 h,LvARMC8基因在凡纳滨对虾血液中的表达量显著降低,但48 h后显著升高,72 h达到最高水平。除副溶血弧菌感染后24 h外的时间点,LvARMC8基因在凡纳滨对虾血液中的表达量均显著升高。研究表明,LvARMC8基因可能参与凡纳滨对虾抗病免疫应答途径。
[Abstract]:In order to study the immune function of Armadillo-containing repeat protein 8 in the process of disease-resistant infection of Litopenaeus vannamei, the full-length c-DNA sequence of ARMC8 gene (LvARMC8,Gen Bank registration number: KX058562) of Litopenaeus vannamei was cloned by RACE-PCR technique for the first time. On-line software was used for bioinformatics analysis. The expression of LvARMC8 gene in different tissues of Penaeus vannamei and infected with white spot syndrome virus and Vibrio parahaemolyticus was analyzed by real-time fluorescence quantitative PCR (Rt-PCR). The results showed that the full length of the LvARMC8 gene was 2917bp. The open reading frame (ORF) was 2046 bp, encoding 681 amino acids, and the 5 'non coding region was 49 bp,3' noncoding region 822 bp.. Predictive analysis showed that the protein encoded by LvARMC8 contained six ARM domains. Homology analysis showed that the similarity between LvARMC8 gene and ARMC8 gene of Nevada termites was the highest (71%). Phylogenetic analysis showed that LvARMC8 and a variety of invertebrate ARMC8 were clustered into one branch, among which, the phylogenetic relationships with insect species, Cephalopheles quinquefasciatus and Citrus spp. Rt-PCR analysis showed that they were closely related to each other. LvARMC8 gene was detected in many tissues of Penaeus vannamei, the highest expression level was found in epidermis and the lowest expression was in eye stalk. After 12 h of WSSV infection, the expression of LvARMC8 gene decreased significantly in the blood of Penaeus vannamei, but increased significantly 48 h after infection, and the expression level of LvARMC8 gene in the blood of Penaeus vannamei was significantly increased after 48 h. 72 h reached the highest level. Except 24 hours after infection with Vibrio parahaemolyticus, the expression of LvARMC8 gene in the blood of Penaeus vannamei increased significantly. The results showed that LvARMC8 gene might be involved in the disease-resistant immune response pathway of Litopenaeus vannamei.
【作者单位】： 广东海洋大学水产学院;南海生物资源开发与利用协同创新中心;
【基金】：广东省自然科学基金—博士启动项目(2014A030310184);广东省自然科学基金—自由申请项目(2016A030313757) 广东普通高校青年创新人才项目(2014KQNCX079) 广东海洋大学优秀青年骨干教师特别资助计划(HDYQ2015004)~~
【分类号】：S917.4
[Abstract]:In order to study the immune function of Armadillo-containing repeat protein 8 in the process of disease-resistant infection of Litopenaeus vannamei, the full-length c-DNA sequence of ARMC8 gene (LvARMC8,Gen Bank registration number: KX058562) of Litopenaeus vannamei was cloned by RACE-PCR technique for the first time. On-line software was used for bioinformatics analysis. The expression of LvARMC8 gene in different tissues of Penaeus vannamei and infected with white spot syndrome virus and Vibrio parahaemolyticus was analyzed by real-time fluorescence quantitative PCR (Rt-PCR). The results showed that the full length of the LvARMC8 gene was 2917bp. The open reading frame (ORF) was 2046 bp, encoding 681 amino acids, and the 5 'non coding region was 49 bp,3' noncoding region 822 bp.. Predictive analysis showed that the protein encoded by LvARMC8 contained six ARM domains. Homology analysis showed that the similarity between LvARMC8 gene and ARMC8 gene of Nevada termites was the highest (71%). Phylogenetic analysis showed that LvARMC8 and a variety of invertebrate ARMC8 were clustered into one branch, among which, the phylogenetic relationships with insect species, Cephalopheles quinquefasciatus and Citrus spp. Rt-PCR analysis showed that they were closely related to each other. LvARMC8 gene was detected in many tissues of Penaeus vannamei, the highest expression level was found in epidermis and the lowest expression was in eye stalk. After 12 h of WSSV infection, the expression of LvARMC8 gene decreased significantly in the blood of Penaeus vannamei, but increased significantly 48 h after infection, and the expression level of LvARMC8 gene in the blood of Penaeus vannamei was significantly increased after 48 h. 72 h reached the highest level. Except 24 hours after infection with Vibrio parahaemolyticus, the expression of LvARMC8 gene in the blood of Penaeus vannamei increased significantly. The results showed that LvARMC8 gene might be involved in the disease-resistant immune response pathway of Litopenaeus vannamei.
【作者单位】： 广东海洋大学水产学院;南海生物资源开发与利用协同创新中心;
【基金】：广东省自然科学基金—博士启动项目(2014A030310184);广东省自然科学基金—自由申请项目(2016A030313757) 广东普通高校青年创新人才项目(2014KQNCX079) 广东海洋大学优秀青年骨干教师特别资助计划(HDYQ2015004)~~
【分类号】：S917.4
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