日本血吸虫表膜结构蛋白Annexin基因克
发布时间：2018-08-11 12:09
【摘要】： 日本血吸虫病是我国一种危害严重的人畜共患寄生虫疾病。由于中间宿主钉螺难以消灭,治疗药物“吡喹酮”不能解决重复感染的难题,并且有可能诱导产生抗药性,因此有必要研制高效、安全的抗血吸虫病疫苗和新治疗药物。血吸虫能够在终末宿主体内生活十几年甚至几十年,一定具有逃避宿主免疫应答的有效方式;血吸虫的体表被膜是由复杂的多层结构组成的合胞体层,它直接受到宿主免疫系统的攻击,血吸虫之所以具有很强的自身保护能力,与其表膜密切相关,因此深入研究血吸虫表膜蛋白及其抗原组成及变化是研究血吸虫与宿主之间相互作用、揭示血吸虫免疫逃避机制的关键。本研究克隆了日本血吸虫表膜蛋白Annexin的编码基因,并对这个基因进行了表达及生物学功能的初步研究。 作者利用已经公布的曼氏血吸虫表膜蛋白Annexin基因序列,在日本血吸虫EST库中搜索到一个相应的EST片段(GeneBank登录号AY812989),根据该EST序列,设计特异性引物,利用PCR技术克隆获得日本血吸虫表膜蛋白Annexin编码基因Sjan。序列分析表明Sjan基因的ORF含1023bp,编码341个氨基酸,理论分子量39.4KD。并成功构建该基因的重组表达质粒pET28a(+)-Sjan,在大肠杆菌BL21(DE3)系统中成功表达,分子量为42.9KD;应用纯化的重组蛋白rSjan免疫BALB/c小鼠,获得了23.27%的减虫率和35.04%的肝组织减卵率。 本文克隆了编码日本血吸虫表膜蛋白Annexin的基因,成功将Sjan在大肠杆菌中表达,并在小鼠中初步评估了该重组蛋白诱导的免疫保护效果。本研究为深入探讨日本血吸虫抵抗宿主免疫系统的攻击、揭示日本血吸虫的免疫逃避机制提供了基础,也为研制高效、安全的抗血吸虫病疫苗和新的治疗药物提供了新思路。
[Abstract]:Schistosomiasis japonicum is a serious zoonotic parasitic disease in China. Because the intermediate host snail is difficult to eliminate, the treatment drug praziquantel can not solve the problem of repeated infection and may induce drug resistance. Therefore, it is necessary to develop an efficient and safe anti-schistosomiasis vaccine and new therapeutic drugs. Schistosoma japonicum can live in the terminal host for more than ten years or even decades, and must have an effective way to escape the host immune response. It is directly attacked by the host immune system. The reason why Schistosoma japonicum has strong self-protection ability is closely related to its epimembrane. Therefore, it is the key to study the interaction between Schistosoma japonicum and its host and reveal the immune escape mechanism of Schistosoma japonicum. In this study, the encoding gene of Schistosoma japonicum surface membrane protein (Annexin) was cloned, and its expression and biological function were studied. Using the published Annexin gene sequence of Schistosoma mansoni surface membrane protein, a corresponding EST fragment (GeneBank accession number AY812989) was found in the Schistosoma japonicum EST library. According to the EST sequence, specific primers were designed. The Annexin encoding gene of Schistosoma japonicum surface membrane protein (Annexin) was cloned by PCR technique. Sequence analysis showed that the ORF of Sjan gene contained 1023 BP, encoding 341 amino acids, and its theoretical molecular weight was 39.4 KD. The recombinant expression plasmid pET28a () -Sjanwas successfully constructed and expressed in Escherichia coli BL21 (DE3) system with a molecular weight of 42.9 KD.The purified recombinant protein rSjan was used to immunize BALB/c mice, and the worm reduction rate of 23.27% and the oocyte reduction rate of 35.04% in liver tissue were obtained. The gene encoding Schistosoma japonicum surface membrane protein (Annexin) was cloned and successfully expressed in Escherichia coli (E.coli). The immune protection induced by the recombinant protein was evaluated in mice. This study provides a basis for further study on the immunity of Schistosoma japonicum against the attack of host immune system, reveals the immune escape mechanism of Schistosoma japonicum, and provides a new idea for the development of efficient and safe anti-schistosomiasis vaccine and new therapeutic drugs.
【学位授予单位】：福建农林大学
【学位级别】：硕士
【学位授予年份】：2009
【分类号】：R392
[Abstract]:Schistosomiasis japonicum is a serious zoonotic parasitic disease in China. Because the intermediate host snail is difficult to eliminate, the treatment drug praziquantel can not solve the problem of repeated infection and may induce drug resistance. Therefore, it is necessary to develop an efficient and safe anti-schistosomiasis vaccine and new therapeutic drugs. Schistosoma japonicum can live in the terminal host for more than ten years or even decades, and must have an effective way to escape the host immune response. It is directly attacked by the host immune system. The reason why Schistosoma japonicum has strong self-protection ability is closely related to its epimembrane. Therefore, it is the key to study the interaction between Schistosoma japonicum and its host and reveal the immune escape mechanism of Schistosoma japonicum. In this study, the encoding gene of Schistosoma japonicum surface membrane protein (Annexin) was cloned, and its expression and biological function were studied. Using the published Annexin gene sequence of Schistosoma mansoni surface membrane protein, a corresponding EST fragment (GeneBank accession number AY812989) was found in the Schistosoma japonicum EST library. According to the EST sequence, specific primers were designed. The Annexin encoding gene of Schistosoma japonicum surface membrane protein (Annexin) was cloned by PCR technique. Sequence analysis showed that the ORF of Sjan gene contained 1023 BP, encoding 341 amino acids, and its theoretical molecular weight was 39.4 KD. The recombinant expression plasmid pET28a () -Sjanwas successfully constructed and expressed in Escherichia coli BL21 (DE3) system with a molecular weight of 42.9 KD.The purified recombinant protein rSjan was used to immunize BALB/c mice, and the worm reduction rate of 23.27% and the oocyte reduction rate of 35.04% in liver tissue were obtained. The gene encoding Schistosoma japonicum surface membrane protein (Annexin) was cloned and successfully expressed in Escherichia coli (E.coli). The immune protection induced by the recombinant protein was evaluated in mice. This study provides a basis for further study on the immunity of Schistosoma japonicum against the attack of host immune system, reveals the immune escape mechanism of Schistosoma japonicum, and provides a new idea for the development of efficient and safe anti-schistosomiasis vaccine and new therapeutic drugs.
【学位授予单位】：福建农林大学
【学位级别】：硕士
【学位授予年份】：2009
【分类号】：R392
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