中国水产科学Issue(6)：1181-1189,9.DOI:10.3724/SP.J.1118.2014.01181
低温胁迫对尼罗罗非鱼水通道蛋白基因（AQP1）表达的影响
Effects of cold stress on Aquaporin gene (AQP1) expression in tilapia Oreochromis niloticus
摘要
Abstract
The full length cDNA of the Aquaporin gene (AQP1) was cloned and sequenced from cold tolerant Nile tila-pia (Oreochromis niloticus) strains developed by successive and directional selective breeding. In addition, the effects of cold stress on AQP1 gene expression levels under low temperature treatments were measured using real-time quan-titative PCR. Sequence analysis showed that Nile tilapia AQP 1 cDNA consists of 1 098 base pairs (bp) and encodes a protein of 261 amino acids that contain a 36 bp 5′untranslated region (UTR), a 783 bp open reading frame (ORF) and a 279 bp 3′UTR. Multiple alignment and homological analysis revealed AQP1 evolutionary conservation among verte-brates and showed that Nile tilapia AQP1 protein shared high levels of amino acid identity with AQP1 from Maylandia zebra (96%), Diplodus sargus (93%), Rhabdosargus sarba (92%), Dicentrarchus labrax (92%), Esox lucius (86%), Anguilla anguilla (83%), Danio rerio (74%), Gallus gallus (63%), Mus musculus (59%) and Homo sapiens (59%). A phylogenetic tree was constructed using the neighbor-joining method with the deduced amino acid sequences of AQP1 among vertebrates. As expected from the sequence alignment, tilapia AQP1 fell into the cluster with Maylandia zebra AQP 1 first, and then clustered with the other teleost species. To investigate the effects of low temperature on AQP1 expression, we examined the expression of AQP1 mRNA at different temperatures (30℃, 25℃, 20℃, 15℃, 10℃) for 48 h using real-time quantitative PCR. There was a gradual decrease in AQP1 mRNA levels in the muscle and liver from 20℃as the water temperature dropped gradually from 30℃ to 10℃. In the muscle of the cold-tolerant tilapia strain, the AQP1 mRNA levels decreased drastically at 20℃and were 22.16 times less at 15℃than at 30℃, and then at 10℃, 107.73 times less down-regulated compared with 30℃. The expression of the AQP1 gene in the control group was 5.38 times less at 15℃than at 30℃, followed by reduced expression at 10℃which induced an 11.30 fold decrease in transcript amounts. Analysis of variance showed that the expression levels of AQP1 in muscle under cold stress were significantly different between the cold-tolerant tilapia strain and the control group (P<0.05). In the liver of the cold-tolerant strain and the control, the AQP1 mRNA expressions were 3.38 and 1.42 fold down-regulated at 15℃, and then at 10℃, 18.85 and 9.01 fold down-regulated when compared with 30℃. This showed that cold stress had no sig-nificant effects on liver AQP1 mRNA expression between the cold-tolerant tilapia strain and the control group (P>0.05). The variation in expression levels suggest that the expression of AQP1 is sensitive to temperature, and the higher cold resistance of the cold-tolerant tilapia strain was closely associated with the drastically down-regulated expression of AQP1 at low temperatures. These results suggest that the AQP1 gene plays an important role in low temperature accli-mation and might be one of the cold tolerance-related genes in Nile tilapia. The present results provide a theoretical basis for studying the molecular mechanism of cold tolerance and fish stress resistance breeding.关键词
尼罗罗非鱼/低温胁迫/水通道蛋白基因/实时荧光定量PCR/基因表达Key words
Nile tilapia (Oreochromis niloticus)/cold stress/Aquaporin gene/real-time quantitative PCR/gene expression分类
农业科技引用本文复制引用
朱华平,刘玉姣,刘志刚,卢迈新,高风英,可小丽,黄樟翰..低温胁迫对尼罗罗非鱼水通道蛋白基因（AQP1）表达的影响[J].中国水产科学,2014,(6):1181-1189,9.基金项目
国家自然科学基金项目(31001108) （31001108）
广东省自然科学基金资助项目(9451038001003595) （9451038001003595）
现代农业产业技术体系资金资助项目(CARS-49) （CARS-49）
国家科技支撑计划专项资金资助项目(2012BAD26B03) （2012BAD26B03）
中国水产科学研究院基本科研业务费资助项目(2012A0404) （2012A0404）
