毒害艾美耳球虫配子体Engam59基因的克隆表达及其表达产物的免疫原性分析

发布时间：2018-07-05 14:57

本文选题：毒害艾美耳球虫 + Engam基因　；参考：《中国兽医科学》2017年12期

【摘要】：提取毒害艾美耳球虫配子体总RNA,应用RT-PCR扩增得到Engam59基因,对其克隆测序并进行序列分析。结果表明,Engam59基因全长为1 473 bp,为一个完整的开放阅读框,编码490个氨基酸,具有1个酪氨酸-丝氨酸富集区和1个脯氨酸-甘氨酸富集区,与柔嫩艾美耳球虫Etgam59基因的同源性为93.4%。选择该基因编码的第211~432位氨基酸间的肽段进行截短表达,获得的重组蛋白大小在27.69 ku左右,主要以包涵体的形式存在。Western-blot分析结果显示,该重组蛋白能被小鼠抗重组蛋白多克隆抗体和毒害艾美耳球虫病鸡康复血清识别,表明该重组蛋白具有很好的免疫原性。本研究结果为进一步探析毒害艾美耳球虫配子体蛋白的功能与研制鸡球虫病基因工程疫苗奠定了基础。
[Abstract]:The total RNAs of gametophyte of Eimeria virulens were extracted. Engam59 gene was amplified by RT-PCR, cloned and sequenced. The results showed that the full length of Engam59 gene was 1 473 BP, which was a complete open reading frame, encoding 490 amino acids, with a tyrosine serine rich region and a proline glycine rich region. The homology between Etgam59 gene and Etgam59 gene of Eimeria tenella is 93. 4%. The peptide fragment of the 21142nd amino acid position encoded by the gene was truncated and expressed. The size of the recombinant protein was about 27.69 ku, which was mainly in the form of inclusion body. The results of Western-blot analysis showed that the recombinant protein could be expressed in the form of inclusion body. The recombinant protein could be recognized by the polyclonal antibody of mouse anti-recombinant protein and the recovered serum of the poisoned Eimeria coccidiosis chicken, which indicated that the recombinant protein had good immunogenicity. The results laid a foundation for further study on the function of gametophyte protein and the development of genetic engineering vaccine for coccidiosis.
【作者单位】：扬州大学兽医学院江苏省高校动物重要疫病与人兽共患病防控协同创新中心;
【基金】：国家重点研发计划项目(2017YFD0501205) 国家自然科学基金项目(31472181,31602039) 江苏省高校自然科学研究面上项目(15KJB230005) 江苏省博士后科研资助计划项目(1601054C) 江苏省高校优势学科建设工程项目
【分类号】：S852.723

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