日本血吸蟲表膜結(jié)構(gòu)蛋白Annexin基因克
發(fā)布時間：2018-08-11 12:09
【摘要】： 日本血吸蟲病是我國一種危害嚴重的人畜共患寄生蟲疾病。由于中間宿主釘螺難以消滅,治療藥物“吡喹酮”不能解決重復(fù)感染的難題,并且有可能誘導(dǎo)產(chǎn)生抗藥性,因此有必要研制高效、安全的抗血吸蟲病疫苗和新治療藥物。血吸蟲能夠在終末宿主體內(nèi)生活十幾年甚至幾十年,一定具有逃避宿主免疫應(yīng)答的有效方式;血吸蟲的體表被膜是由復(fù)雜的多層結(jié)構(gòu)組成的合胞體層,它直接受到宿主免疫系統(tǒng)的攻擊,血吸蟲之所以具有很強的自身保護能力,與其表膜密切相關(guān),因此深入研究血吸蟲表膜蛋白及其抗原組成及變化是研究血吸蟲與宿主之間相互作用、揭示血吸蟲免疫逃避機制的關(guān)鍵。本研究克隆了日本血吸蟲表膜蛋白Annexin的編碼基因,并對這個基因進行了表達及生物學(xué)功能的初步研究。 作者利用已經(jīng)公布的曼氏血吸蟲表膜蛋白Annexin基因序列,在日本血吸蟲EST庫中搜索到一個相應(yīng)的EST片段(GeneBank登錄號AY812989),根據(jù)該EST序列,設(shè)計特異性引物,利用PCR技術(shù)克隆獲得日本血吸蟲表膜蛋白Annexin編碼基因Sjan。序列分析表明Sjan基因的ORF含1023bp,編碼341個氨基酸,理論分子量39.4KD。并成功構(gòu)建該基因的重組表達質(zhì)粒pET28a(+)-Sjan,在大腸桿菌BL21(DE3)系統(tǒng)中成功表達,分子量為42.9KD;應(yīng)用純化的重組蛋白rSjan免疫BALB/c小鼠,獲得了23.27%的減蟲率和35.04%的肝組織減卵率。 本文克隆了編碼日本血吸蟲表膜蛋白Annexin的基因,成功將Sjan在大腸桿菌中表達,并在小鼠中初步評估了該重組蛋白誘導(dǎo)的免疫保護效果。本研究為深入探討日本血吸蟲抵抗宿主免疫系統(tǒng)的攻擊、揭示日本血吸蟲的免疫逃避機制提供了基礎(chǔ),也為研制高效、安全的抗血吸蟲病疫苗和新的治療藥物提供了新思路。
[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.
【學(xué)位授予單位】：福建農(nóng)林大學(xué)
【學(xué)位級別】：碩士
【學(xué)位授予年份】：2009
【分類號】：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.
【學(xué)位授予單位】：福建農(nóng)林大學(xué)
【學(xué)位級別】：碩士
【學(xué)位授予年份】：2009
【分類號】：R392
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