摘要
通过同源克隆策略并结合cDNA末端快速扩增 (RACE)技术 ,克隆到了编码家蝇攻击素 (Attacin)基因的全长cDNA序列 (GenBank登陆号 :AY4 6 0 10 6 ) ,并对相应的氨基酸序列进行了序列比对 ,对系统关系等进行了生物信息学分析。家蝇Attacin的cDNA全长 778bp ,其中包括 6 2 7bp的开放式读码框 (ORF) ,4 4bp的 5′末端非编码区 (5′UTR)和 10 7bp的 3′末端非编码区 (3′UTR) ,相应的蛋白产物含 2 0 8个氨基酸 ,与其他双翅目昆虫的Attacin具有很高的相似性 ,约 5 0 %～ 70 %。以邻接法 (Neighbor Joining ,NJ)所构建的系统关系表明 ,家蝇的Attacin与其他双翅目昆虫的Attacin起源于共同的祖先。应用半定量RT PCR的方法 ,研究家蝇幼虫在受到外源刺激后Attacin基因的表达 ,结果表明 ,在家蝇幼虫体内Attacin基因呈诱导型表达 。
The antimicrobial peptides of insect are the main components of their non-specific immune system,and play a major role in the defense against the foreign disease-related microbes.In this report,a full length cDNA of Attacin,an insect antimicrobial peptide was cloned from housefly (Musca domestica) by homology cloning approach in combine with 3′ and 5′ RACE.Sequence analysis and phylogenetical study showed that this cDNA contained 778 nucleotides,with a 627 bp open reading frame (ORF) flanked with a 44 bp 5′UTR and a 107 bp 3′UTR.The encoded 208 amino acids housefly Attacin shared a high similarity of 50%～70% with that of the other dipterous insects.In addition,the phylogenetical analysis also indicated that the Attacin from housefly was in the same branch with those of other species,suggesting that they come from the same ancestor.The expression of Attacin transcript was measured by semi-quantitive RT-PCR.The results demonstrated that the expression of housefly Attacin is inducible,and that the level varies with the time of induction and the kinds of pathogens.
基金
天津市自然科学基金资助项目 (项目编号 :0 3 3 60 4911)~~
关键词
家蝇
攻击素
同源克隆
RT—PCR
housefly (Musca domestica)
Attacin
homology cloning
RT-PCR
