CAJ | 학술논문

转铁蛋白是呼吸链的关键因子, 参与体液和免疫系统的调节, 具有抗菌抗病毒等功能.研究从嗜水气单胞菌诱导的中华鳖肝脏SMART cDNA文库中,分离克隆了中华鳖转铁蛋白基因cDNA和gDNA全序列,对其序列特征、组织表达及原核表达进行了分析研究.结果显示, 中华鳖转铁蛋白对应全长cDNA的基因组DNA全长为19100 bp,由17个外显子和16个内含子组成; cDNA全长为2359 bp,开放阅读框为 2094 bp,编码697个氨基酸;转铁蛋白的分子量为76.6 kD,包含2个糖基化位点,存在4个二硫键形成区域的缺失,两个同源结构域的相似性为 37%, 信号肽位于第 1—19 位点.荧光定量 PCR 结果显示, 正常组中华鳖转铁蛋白基因在肝脏中的表达量最大; 以嗜水气单胞菌刺激后, 其在心、肝、脾和肾脏组织中的表达, 均有一个上调后减少的趋势.此外,实验还进行了重组转铁蛋白原核表达和Western blot检测鉴定,为深入研究中华鳖转铁蛋白功能的提供了基础数据.
전철단백시호흡련적관건인자, 삼여체액화면역계통적조절, 구유항균항병독등공능.연구종기수기단포균유도적중화별간장SMART cDNA문고중,분리극륭료중화별전철단백기인cDNA화gDNA전서렬,대기서렬특정、조직표체급원핵표체진행료분석연구.결과현시, 중화별전철단백대응전장cDNA적기인조DNA전장위19100 bp,유17개외현자화16개내함자조성; cDNA전장위2359 bp,개방열독광위 2094 bp,편마697개안기산;전철단백적분자량위76.6 kD,포함2개당기화위점,존재4개이류건형성구역적결실,량개동원결구역적상사성위 37%, 신호태위우제 1—19 위점.형광정량 PCR 결과현시, 정상조중화별전철단백기인재간장중적표체량최대; 이기수기단포균자격후, 기재심、간、비화신장조직중적표체, 균유일개상조후감소적추세.차외,실험환진행료중조전철단백원핵표체화Western blot검측감정,위심입연구중화별전철단백공능적제공료기출수거.
Transferrin has a variety of biological functions such as iron transportation, anti-microbial, and immu-noregulation. In this study, we isolated the full length cDNA of transferrin of Chinese soft shell turtle (Pelodiscus sinensis) from the constructed liver cDNA library. The cDNA sequence consisted of 2359 bp, including an open reading frame (ORF) of 2094 bp which could encode a 674-aa peptide. The genomic DNA of transferrin consisted of 19100 bp and contained 17 exons and 16 introns, with a structure similar to that of other vertebrates. Using the online software we predicted the molecular characteristics of transferrin. The results indicated that the N-lobe of transferrin had a signal peptide of 19 amino acids; this gene was composed of two similar domains with a similarity of 37%. There were 2 N-linked glycosylation sites in the N-lobe, and 11 disulfide bonding formation areas in amino acids peptide. The quan-titative PCR results showed that the expression of transferrin was the highest in liver in the normal group. The results also showed that after the infection withAeromonas hydrophila the expression of this gene was first increased and then decreased in liver, spleen, kidney and heart, and the increase was particularly high in liver and spleen. Moreover, we applied western-blot to testify the prokaryotic expression of transferrin of Chinese soft shell turtle. This study could provide insights into the biological functions of transferrin in non-specific immune responses.