| 摘要: |
| 【目的】克隆鸡柔嫩艾美耳球虫SO7基因并进行原核表达，检测表达产物的免疫原性，为鸡球虫基因疫苗的制备奠定基础。 【方法】以纯化的柔嫩艾美耳球虫杨凌（Eimeria tenella YL）株孢子化卵囊的总RNA为模板，用RT-PCR方法扩增出E.tenella YL 株的SO7基因，对其测序后进行序列分析。将SO7基因克隆到表达载体pET-32a中，构建pET-SO7重组质粒，并在大肠埃希菌BL21中诱导表达出重组蛋白，将重组蛋白纯化后免疫雏鸡，评价其免疫效果。【结果】鸡柔嫩艾美耳球虫SO7基因序列的开放阅读框（ORF）为651个碱基，共编码217个氨基酸。E.tenella YL株SO7基因与E.tenella LS18株、E.tenella BJ株、E.tenella GD株SO7基因ORF区序列相似性分别为99.1%，98.6%和99.1%，其氨基酸序列相似性分别为99.1%，98.6%和99.1%。SDS-PAGE分析表明，表达产物成功地表达出了分子质量为45 ku的融合蛋白。各种抗球虫指标的测定结果显示，该蛋白对球虫感染鸡体具有一定的保护性。【结论】SO7基因具有很强的保守性，各虫株之间差异不大；重组蛋白具有一定的保护效果，适合用来制备基因疫苗。 |
| 关键词: SO7基因 柔嫩艾美耳球虫杨凌株 克隆表达 保护性试验 |
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| 基金项目:陕西省农业攻关项目（2008K02-06）；985工程科技创新平台项目“畜禽养殖与重大疾病防治”（Z101020001） |
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| Cloning and expression of E.tenella YL strain SO7 gene and protective experiments |
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| Abstract: |
| 【Objective】The research studied cloning and expression of E.tenella YL strain SO7 gene.【Method】E.tenella SO7 gene was cloned from E.tenella sporulated oocysts sporozoites by RT-PCR.The recombinant expression plasmid pET-SO7 was constructed and expressed in BL21 strain of E.coli.The product was analysed by SDS-PAGE.Then,recombinant proteins were used in chickens infected with E.tenella YL strain to judge the protective rate of the recombined proteins.【Result】Sequence analysis suggested that the ORF of SO7 gene consisted 651 nucleotides and encoded 217 amino acids.Compared with the E.tenella LS18,E.tenella BJ and E.tenella GD strain,the homology of nucleotides was 99.1%,98.6% and 99.1%,the homology of amino acid 99.1%,98.6% and 99.1%.SDS-PAGE showed that the fusion protein was successfully expressed and the molecular weight of the protein was 45 ku.Protective experiments showed that the recombinant proteins induced certain anticoccidial immune protection.【Conclusion】SO7gene was highly reserved,and little difference existed between the various strains.The recombinant proteins had certain anticoccidial protection,suitable for preparation of DNA vaccine. |
| Key words: SO7 gene Eimeria tenella YL strain cloning and expression protective experiment |
