日本蟾蜍皮肤claudin-4 cDNA的克隆与序列分析

摘要

为研究日本蟾蜍Bufo japonicus formosus皮肤中多肽类有效成分,通过菌落聚合酶链式反应(PCR)对日本蟾蜍皮肤cDNA质粒文库进行筛选,克隆到2条claudin-4全长cDNA序列(GenBank登录号为JX481975和JX481976),并对它们进行生物信息学分析.日本蟾蜍claudin-4的2条cDNA全长分别为1 707 bp和1 697 bp,开放阅读框均为630 bp,其中虽有5个碱基不同,但编码的209个氨基酸完全相同.两者的5'端非翻译区域序列长度不同,前者为50 bp,后者为41 bp,3'端非翻译区域则分别为1 027 bp和1 026 bp.氨基酸序列同源性分析显示,日本蟾蜍claudin-4与非洲爪蟾Xenopus laevis的同源性最高,为88％,与其他7种动物的同源性介于65％～72％.克隆到2条日本蟾蜍claudin-4基因的cDNA序列,为今后研究日本蟾蜍Claudin-4生物学功能奠定了基础.%To clone the genes encoding the bioactive polypeptides included in Bufo skin and its secretions,the skin plasmid cDNA library of the adult Japanese toad Bufo japonicus formosus was screened by colony DNA polymerase chain reaction followed by a homologous analysis.Results revealed two full length cDNAs of claudin-4.The transcripts were 1 707 bp and 1 697 bp (GenBank:JX481975 and JX481976,respectively),both containing an open reading frame (ORF) of 630 bp.The same polypeptides consisting of 209 amino acid residues were encoded,although five nucleotides were different in the ORF.The 5' untranslated region (UTR) for the two cDNAs was 50 bp and 41 bp,in that order,while the 3' UTR was 1 027 bp and 1 026 bp,respectively.The homologous analysis indicated that Claudin-4 in B.japonicus formosus had an 88％ similarity with Xenopus laevis and 65％-72％ similarity with other species including Taeniopygia guttata,Anolis carolinensis,Danio rerio,Ornithorhynchus anatinus,Macaca mulatta,Mus musculus,Homo sapiens.This study will become the basis for further studies on biological functions of Claudin-4 in B.japonicus formosus and the drug development for tumor theraputics.