中国林蛙皮肤抗菌肽基因的cDNA克隆  

cDNA Cloning of the Antibacterial Peptide of the Rana Temporaria Chensinensis David Skin

作　　者：乔红伟[1] 韩俊友[1] 赵瑞利[1] 郑伟[2] 韩艳[1] 韩文瑜[1] 

机构地区：[1]吉林大学畜牧兽医学院,吉林长春130062 [2]吉林省水产研究院,吉林长春130062

出　　处：《中国兽医学报》2006年第6期646-648,共3页Chinese Journal of Veterinary Science

基　　金：国家自然科学基金项目(30340087);吉林省科技厅应用基础项目

摘　　要：抗菌蛋白蛙皮抗菌肽,是两栖类动物中存在的天然免疫物质,在蛙类中仍然保持着一定的活性。我们用提取中国林蛙皮肤总RNA,逆转录合成cDNA,PCR扩增得到DNA片段,并与GenBank上登陆的其他蛙类的皮肤抗菌肽序列进行比较,发现此序列是一个新的蛙皮肤抗菌肽序列。PCR产物与pMD18-T连接,得到阳性重组体pMD18-RTCD,将阳性重组体进行测序,分析蛙皮素的氨基酸序列,显示它属于蛙皮抗菌肽家族。此外,中国林蛙抗菌肽氨基酸序列包含C-末端残基,同时具有两栖类动物抗菌肽的特征。这些信息对将来研究两栖类动物的天然免疫的分子机制具有重要作用,为应用基因工程技术对我国林蛙抗菌肽基因进行克隆表达奠定了基础。Antibacterial peptide of rana skin, an antibacterial protein, has been implicated in innate immunity m invertebrates. We cloned the Rana ternporaria chensinensis David skin cDNA using a PCR strategy and detected its activity in rana using a antibacterial protein assay against Micrococcus luteus. The cloning was based on a report of GenBank. The deduced amino acid sequence resulted in 53 amino with 46% identity to amphibia. RT-PCR was used to detect peptide mRNA in Rana temporaria chensinensis David. PCR products with T-4 linkagemse and pMD18-T vector. A positive recombinant clone pMD18-Cla obtained. Sequencing and anslysis of the amino acid sequence showed that the sequence contained extra residues at its C-type family of antibacterial peptide. Furthermore,the antibacterial peptide amino acid sequence contained extra residues at its C-terminus,which are characteristic of marine inverterates. This information will be useful in future studies on the molecular mechanisms of immunity in marine invertebrates,and provides the base for preparing the skin antibacterial peptide of the Rana temporaria chensinensis David with genetic engineering method.

关 键 词：蛙皮抗菌肽基因 中国林蛙 PCR克隆 

分 类 号：S852.4[农业科学—基础兽医学]