细粒棘球绦虫EgM123基因序列分析及EgM家族基因在虫体不同发育阶段的差异表达分析

发布时间：2018-06-19 21:24

本文选题：细粒棘球绦虫 + EgM蛋白　；参考：《动物医学进展》2017年10期

【摘要】：对细粒棘球绦虫EgM123基因进行分子克隆和生物信息学分析,同时对EgM家族基因在虫体不同发育阶段的mRNA转录水平进行初步研究。采集绵羊肝脏包囊中原头蚴,提取总RNA,通过RT-PCR获取EgM123纯化的PCR产物,克隆到pGEM-T载体并测序,对测序结果进行结构和功能的预测分析。以Actin基因为内参基因,采用SYBR Green real-time RT-PCR技术,对不同发育阶段,即其原头蚴和成虫的EgM4、EgM9、EgM123的表达量进行检测。结果表明,EgM123基因ORF为594bp,编码197个氨基酸,蛋白分子质量为21.959ku,等电点为7.94。EgM123蛋白无信号肽、跨膜螺旋区和卷曲螺旋结构,是一种亲水性的可溶性蛋白质,存在23个磷酸化位点。根据亚细胞定位预测显示,EgM123蛋白主要位于细胞核内。二级结构预测含α-螺旋9.64%、β-折叠22.84%、无规则卷曲62.94%。EgM123氨基酸序列中有6个潜在的抗原表位。RT-qPCR结果显示,成虫阶段EgM家族基因表达量极显著高于原头蚴阶段。在发育的同一阶段,EgM123基因极显著高于EgM4、EgM9基因的表达量。通过EgM家族基因在虫体不同发育阶段的mRNA转录水平,EgM123基因在虫体成虫阶段高表达,以期为筛选细粒棘球绦虫终末宿主疫苗候选基因提供理论依据。
[Abstract]:The EgM123 gene of Echinococcus granulosus was cloned by molecular cloning and bioinformatics analysis, and the mRNA transcription level of EgM family genes at different stages of development of Echinococcus granulosus was studied. The purified PCR product of EgM123 was obtained by RT-PCR and cloned into pGEM-T vector and sequenced. The structure and function of the sequencing results were predicted and analyzed. Using Actin gene as internal reference gene, the expression of EgM4, EgM9, EgM123 was detected by SYBR Green real-time RT-PCR in different developmental stages, I. e., egg M4, EgM9, EgM123. The results showed that the ORF of EgM123 gene was 594bp, encoding 197 amino acids, protein molecular weight was 21.959ku.Isoelectric point was 7.94.EgM123 protein without signal peptide, transmembrane helical region and curly helical structure, it was a hydrophilic soluble protein with 23 phosphorylation sites. According to the subcellular localization prediction, the egg M123 protein was mainly located in the nucleus. The prediction of secondary structure includes 伪 -helix 9.64, 尾 -folding 22.84b and irregular curl 62.94. EgM123 amino acid sequence. The results of RT-qPCR showed that the expression of EgM family genes in adult stage was significantly higher than that in proto cercariae stage. At the same stage of development, the expression of EgM123 gene was significantly higher than that of EgM4 EgM9 gene. In order to provide theoretical basis for screening candidate genes of Echinococcus granulosus final host vaccine, EgM family genes were overexpressed at different stages of mRNA transcription of Echinococcus granulosus at different stages of development of Echinococcus granulosus (Echinococcus granulosus).
【作者单位】：新疆农业大学动物医学学院;省部共建绵羊遗传改良和健康养殖国家重点实验室;
【基金】：国家自然科学基金项目(31360623) 研究生科研创新项目(XJAUGRI2015017)
【分类号】：S852.734

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