羊泰勒虫核苷酸合成代谢途径初步分析及OPRT蛋白的免疫保护试验

发布时间：2018-03-02 16:36

本文选题：羊泰勒虫　切入点：蛋白组OPRT　出处：《中国农业科学院》2015年硕士论文　论文类型：学位论文

【摘要】：羊泰勒虫病是由媒介蜱传播的泰勒科泰勒属的原虫寄生于绵羊和山羊的巨噬细胞、淋巴细胞和红细胞内而引起的疾病的总称,它给全球养羊业带来了巨大的经济损失。本研究运用生物信息学初步分析了羊泰勒虫蛋白组的核酸合成代谢路径,进行了乳清酸磷酸核糖转移酶(OPRT)基因的克隆表达,以及rOPRT的绵羊保护性效果评价。其研究结果如下:1.对尤氏泰勒虫、吕氏泰勒虫两个种的六个地方株的蛋白组进行核苷酸代谢途径初步分析,用COG的聚类分析结果表明,与核苷酸代谢相关的蛋白有5个,其中4个参与从头合成途径,只有一个参与补救途径。其中乳清酸磷酸核糖转移酶、腺苷酸激酶分别为从头合成途径、补救途径的关键酶,具有重要的研究意义。2.克隆和表达了尤氏泰勒虫oprt基因的功能区片段,并对重组蛋白rOPRT的反应原性进行分析。结果表明oprt基因的表达产物为包涵体蛋白,rOPRT除与尤氏泰勒虫阳性血清反应外,也与吕氏泰勒虫和绵羊泰勒虫阳性血清发生交叉反应。证实oprt同源基因也存在于吕氏泰勒虫和绵羊泰勒虫。3.动物保护性实验完成了rOPRT对绵羊免遭尤氏泰勒虫感染的保护效果评估。结果表明,对照组绵羊发烧持续时间明显长于免疫组,与对照组相比,免疫组绵羊带虫期明显缩短。对照组实验绵羊分别在攻虫血后第25天、26天、29天出现高虫血症,而免疫组则分别出现在第26天、32天和33天,明显比对照组出现的晚。血红蛋白含量测定的结果显示,对照组的血红蛋白含量(Hb)显著下降,而实验组Hb变化较小。同时红细胞计数和临床症状观察,可以看出对照组贫血严重且持续时间长。4.用ELISA对2012年5月到2013年6月份期间采自中国21个省份、47个地区的8至20月龄绵羊和山羊的2351份血清进行了检测,对我国羊泰勒虫病的流行情况进行调查,结果表明全国各地血清学阳性率存在明显差异,如甘肃省羊的泰勒虫病血清学阳性率最高,为76%,而河北省血清学阳性率最低,未检测到羊泰勒虫感染。
[Abstract]:Sheep Taylor's disease is a disease caused by the parasites of the genus Thaleridae, which is spread by vector ticks, in macrophages, lymphocytes and red blood cells of sheep and goats. It has brought great economic losses to the global sheep industry. In this study, bioinformatics was used to analyze the nucleic acid biosynthesis pathway of sheep Taylor's proteome, and to clone and express the nucleic acid transcriptase (OPRT) gene of whey acid phosphate ribonucleosyltransferase (OPRT). The results of this study are as follows: 1. The nucleotide metabolic pathways of six local strains of Taylor Eurex and Taylor lui were preliminarily analyzed, and the results of COG cluster analysis showed that, There are five proteins related to nucleotide metabolism, four of which are involved in the ab initio pathway, and only one is involved in the rescue pathway, in which whey acid phosphate ribosyltransferase and adenylate kinase are the key enzymes in the ab initio synthesis pathway, respectively. Cloning and expression of the functional region of the oprt gene of Taylor Eurex. The reactivity of the recombinant protein rOPRT was analyzed. The results showed that the expression product of oprt gene was the inclusion body protein rOPRT, which reacted with the positive serum of Taylor's parasite. It was confirmed that oprt homologous gene was also present in Taylor lui and Taylor's sheep. 3. Animal protective experiment was completed to protect sheep from Taylor lucifera infection. The results show that, The duration of fever in the control group was significantly longer than that in the immunized group, and compared with that in the control group, the period of infection in the control group was significantly shorter than that in the control group. The experimental sheep in the control group developed hyperwormemia on the 25th day, the 26th day and the 29th day, respectively. However, the immunized group appeared on the 26th day, 32 days and 33 days, respectively, which was obviously later than that of the control group. The results of hemoglobin content test showed that the hemoglobin content of the control group decreased significantly. The HB of the experimental group was small, and the red blood cell count and clinical symptoms were observed. It can be seen that anemia in the control group is severe and lasting for a long time. ELISA was used to detect 2,351 serum samples of sheep and goats aged 8 to 20 months collected from 21 provinces and 47 regions of China from May 2012 to June 2013. The prevalence of Taylor's disease in sheep in China was investigated. The results showed that there were significant differences in seropositive rate between different parts of the country. For example, the seropositive rate of Taylor's disease in sheep in Gansu Province was the highest (76%), while the seropositive rate in Hebei Province was the lowest. No Taylor's infection was detected in sheep.
【学位授予单位】：中国农业科学院
【学位级别】：硕士
【学位授予年份】：2015
【分类号】：S852.7

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