鳜胃蛋白酶原基因cNA全长的克隆与序列分析.docVIP

鳜胃蛋白酶原基因cDNA全长的克隆与序列分析 200090） 关键词：鳜；胃蛋白酶原；cDNA末端快速扩增；克隆；序列分析 Cloning and sequencing of the full-length cDNA of pepsinogen gene from the Mandarin fish，Siniperca chuatsi ZHAO Jin-Liang (Key Laboratory of Aquatic Genetic Resources and Aquacultural Ecosystem Certificated by the Ministry of Agriculture, Shanghai Fisheries University, Shanghai 200090, China) Abstract: Pepsinogen is one kind of gastric digestive proteinases belongling to a family of aspartic proteinases, which is synthesized in the gastric mucosa of vertebrates and converted to pepsins in the acidic environment of gastric juice. The mandarin fish (Siniperca chuatsi) is a typical carnivorous fish which only prey on small live fishes after hatching. In order to understand the molecular characters of pepsinogen in fish, the full-length cDNA of pepsinogen gene of S. chuatsi was cloned by means of RT-PCR and rapid amplification of cDNA ends (RACE) and sequenced. The result reveals a 1367 bp cDNA full-length sequence containing 43 bp 5′-untranslated region, 187 bp 3′-untranslated region and 1137 bp open reading frame (ORF), which encodes 378 amino acids with characteristic signal peptides and activation peptides. The pepsinogen of S. chuatsi also has highly conserved amino acid residues essential for catalytic activity and conformational maintenance, which are two essential aspartyl residues in the active site and six cysteine residues involved in forming three disulphide bonds. The homology of amino acid sequences between pepsinogen of S. chuatsi and other vertebrate pepsinogens ranges from 59.9% to 91.2%, which indicates the pepsinogen gene is highly conserved in evolution. The NJ phylogenetic tree of vertebrates from the amino acids sequences of pepsinogen showed all fish were clustered as a group, the pepsinogen of S. chuatsi was the most close to pepsinogen A form Ⅱa of winter flounder. The successful cloning of pepsinogen gene from S. chuatsi not only lays the foundation for further study of its tem