猪瘟病毒石门株动物传代脾毒与组织培养细胞毒E2基因序列的分析     

魏中锋  张彦明  孙裴 《西北农林科技大学学报(自然科学版)》2005,33(6):45-48

根据已发表的猪瘟病毒基因组序列,设计合成了2对引物,以脾毒和细胞毒总RNA为模板,利用反转录聚合酶链式反应(RT-PCR)、套式聚合酶链式反应(nPCR)及测序技术,对多次动物传代和致细胞病变的猪瘟病毒主要囊膜糖蛋白E2基因的核苷酸序列进行了测定,用DNAstar软件比较分析了E2基因的核苷酸和氨基酸序列的同源性。结果发现,脾毒和标准石门株核苷酸序列的同源性为99.4%,氨基酸序列的同源性为99.0%;细胞毒与标准石门株核苷酸序列的同源性为98.3%,氨基酸序列的同源性为96.7%。由此说明,猪瘟病毒经猪体多次传代和组织培养致细胞病变后,均能保持遗传的稳定性。  相似文献   

猪瘟病毒石门株E2基因4个抗原结构域的原核表达   总被引：1，自引：1，他引：1  

徐璐  范学政  王琴  蒋春燕  宁宜宝  王泰健 《中国农业科学》2006,39(4):814-818

【目的】对猪瘟病毒石门株E2基因进行原核表达，以期获得可溶性表达产物，为检测猪瘟抗体的ELISA试剂盒的研制奠定基础。【方法】用PCR技术扩增了重组S21质粒载体上的猪瘟病毒石门株E2基因的4个主要抗原结构域ABCD，A1A2，B和C。分别将4个片段克隆于pMAL－p2X载体中，经PCR、双酶切和测序鉴定，E2基因的4个主要抗原结构域片段的位置、大小和读码框均正确。将4个片段分别转化到表达菌TB1、BL21、BL21－CodonPlus(DE3)-RP和BL21(DE3)中，共得到16株重组表达菌，用IPTG进行诱导表达，对表达产物进行SDS-PAGE电泳和免疫印迹分析。【结果】E2基因的4个主要抗原结构域均可在这4种表达菌中表达，但以BL21－CodonPlus(DE3)-RP的表达效果最好，可表达出可溶性并且产量较高的目的蛋白。免疫印迹结果表明，表达的目的蛋白可以被猪瘟阳性血清和针对E2蛋白的单克隆抗体所识别。【结论】只表达目的基因的抗原结构域，可以缩短表达片段的长度，有利于获得可溶性目的蛋白，并且具有良好的血清学反应的特异性。  相似文献   

猪Jiv基因的克隆表达与多克隆抗体制备     

刘伟  郭抗抗  林鸷 《西北农林科技大学学报(自然科学版)》2015,43(2):7-13,22

【目的】克隆猪Jiv蛋白核心区的2 112bp的基因片段,构建其原核表达载体后进行蛋白的表达和纯化,制备抗猪Jiv蛋白的多克隆抗体。【方法】利用RT-PCR方法,从猪脾脏组织RNA中扩增得到猪Jiv基因,将其克隆到pMD19-T载体并测序,以此为基础构建原核表达载体pET32a-Jiv,转化E.coli Rosetta(DE3)感受态细胞中,利用IPTG诱导表达,获得大量包涵体形式的猪Jiv融合蛋白。通过镍离子螯合层析柱对表达的目的蛋白进行纯化,对纯化后的包涵体蛋白进行透析复性,以每只新西兰白兔400μg蛋白的初次免疫剂量和500μg蛋白的加强免疫剂量进行免疫,共免疫5次,采集血清,采用间接ELISA法检测抗体效价达到一定水平后,收集抗体,利用Western blot检测抗体特异性。【结果】克隆得到长度为2 112bp的猪Jiv基因片段;构建的pET32a-Jiv在E.coli Rosetta(DE3)感受态细胞中经诱导后高效表达,SDS-PAGE结果显示,纯化得到分子质量约为95ku的猪Jiv融合蛋白;测得的猪Jiv蛋白多克隆抗体效价达1∶6 400;Western blot检测结果证实,所获得抗体能够有效地应用于猪Jiv蛋白抗原的检测。【结论】成功地克隆了猪Jiv基因,制备了抗猪Jiv蛋白的多克隆抗体。  相似文献   

浅谈猪场高热病的防控措施和要点     

赖厚辉  王培樟  曾国华 《北京农业》2011,(18):100

猪场高热病最科学的防控措施就是要坚持"预防为主,防重于治"的原则,正确科学地制定合理的免疫程序。  相似文献   

抗猪瘟病毒单克隆抗体的研究——Ⅱ抗猪瘟兔化弱毒单克隆抗体杂交瘤细胞株的建立     

唐玉新  周作红  王亚鸣  沈秋姑  徐兰芳  王承浩 《江西农业大学学报》1990,(3)

用聚乙二醇沉淀浓缩纯化猪瘟兔化弱毒中国株(SFV-C)免疫 BALB/C 小鼠,取其脾脏,制备淋巴细胞,与 FO 骨髓瘤细胞融合,经 EliSA 检测和有限稀释法克隆化,最后筛选出5株对SFV 的单克隆抗体杂交瘤细胞.所制成的腹水抗体,一株只对 SFV-C 发生特异性反应,四株与SFV-S,SFV-F 及 BVDV oregon 发生微弱的交叉反应.细胞上清液的 EliSA 效价为1:800～1600,腹水效价为1:10~6～10~7.  相似文献   

Social-ethical issues concerning the control strategy of animal diseases in the European Union: A survey   总被引：1，自引：0，他引：1  

Nina E. Cohen  Marcel A.P.M. van  Asseldonk  Elsbeth N. Stassen 《Agriculture and Human Values》2007,24(4):499-510

In 2004 a survey was conducted in the member states of the European Union designed to gain greater insight into the views on control strategies for foot and mouth disease, classical swine fever, and avian influenza with respect to the epidemiological, economic and social-ethical consequences of each of these animal diseases. This article presents the results of the social-ethical survey. A selection of stakeholders from each member state was asked to prioritize issues for the prevention and control of these diseases. A majority of stakeholders chose preventive measures as the preferred issue. An analysis was done to determine whether there were differences in views expressed by stakeholders from member states with a history of recent epidemics and ones without such a history, and whether there were regional differences. There were no differences between member states with or without a history of recent epidemics. There were indeed regional differences between the priority orders from Northern and Southern Europe on the one hand, and from Eastern Europe on the other. Nina E. Cohen is a biologist and is a researcher at the Wageningen University. She is specialized in societal and ethical issues in human–animal relationships. Her current research is focused on the social-ethical issues concerning the prevention and control of foot and mouth disease, classical swine fever and avian influenza. Marcel A.P.M. van Asseldonk has studied animal science. Currently he works at the Institute for Risk Management in Agriculture (IRMA) of the Wageningen University. He is specialized in the design and pricing of insurance policies and animal health funds for the main livestock epidemics. Elsbeth N. Stassen is a veterinarian and professor of Animals and Society at the Wageningen University. Elsbeth Stassen is specialized in animal health, animal welfare and human–animal relationships. She was a member of a governmental welfare committee during the avian influenza epidemic in the Netherlands in 2003.  相似文献   

The Lapinized Chinese Strain Vaccine Against Classical Swine Fever Virus： A Retrospective Review Spanning Half A Century   总被引：2，自引：0，他引：2  

QIU Hua-ji SHEN Rong-xian TONG Guang-zhi 《中国农业科学(英文版)》2006,5(1):1-14

Classical swine fever （CSF）, a list A disease of Office International des Epizooties, is caused by classical swine fever virus （CSFV） belonging to the Flaviviridae family. The well-known lapinized Chinese strain of CSFV, also known as C-strain, was developed in China in the mid-1950s. In the past half a century, the vaccine has been proved to be safe and immunogenic in pigs of essentially any age. It is of high efficacy, providing immunized animals with broad-spectrum, sometimes lifelong, protection, which is contributed by both cell-mediated immunity and humoral immunity, against essentially all genotypes or subgenotypes of the virus. The maternal antibodies derived from immunized sows can confer solid protection of their offspring from disease; however, they have been proved to inhibit the successful active immunization of C-strain vaccine. The complete genomes of C-strain and dozens of established or field strains have been sequenced and annotated. Recently, the reverse genetics system of C-strain has been developed, resulting in several C- strain-derived candidate marker vaccines. Many countries manage to control or even eradicate CSF with the aid of mass vaccination with C-strain. in spite of these efforts, the eradication of the disease worldwide remains a big challenge and needs to go a long way, and provably still resorts to genetically modified C-strain vaccine. The authors present an overview of the characteristics of the vaccine, which has stood the test of half a century.  相似文献   

内蒙古猪瘟流行病学调查及猪瘟病毒的分离   总被引：3，自引：0，他引：3  

郝永清  王潇  张爱荣  赵和平  刘长春  张秉升 《内蒙古农业大学学报(自然科学版)》2004,25(4):1-4

对内蒙古部分地区猪场进行了猪瘟流行病学调查，发现我区在较大范围内以非典型的猪瘟为主，发病多见于3月龄以下的仔猪，且以局部流行为主，母猪多呈隐性感染，部分母猪表现出繁殖障碍；个别地区仍有发病率和致死率较高的急性典型猪瘟，发病猪场不同程度上存在免疫程序不合理和免疫抑制性疾病。在此基础上对疑似病例进行了病理学检测和兔体交互免疫试验进行确诊，并对猪瘟野毒进行了分离。本研究不仅为内蒙古地区进一步作好猪瘟的防制提供了理论依据，而且为猪瘟野毒分子生物学的研究打下了基础。  相似文献   

The Lapinized Chinese Strain Vaccine Against Classical Swine Fever Virus: A Retrospective Review Spanning Half A Century     

QIU Hua-ji  SHEN Rong-xian  TONG Guang-zhi 《中国农业科学(英文版)》2006,5(1)

  相似文献   

The lesional changes and pathogenesis in the kidney in african swine fever     

J. Hervás  J. C. Gómez-Villamandos  A. Méndez  L. Carrasco  M. A. Sierra 《Veterinary research communications》1996,20(3):285-299

African swine fever is a viral haemorrhagic disease of pigs which has been used as a model for the study of viral haemorrhagic diseases in man. The acute course of the disease is characterized by acute proliferative glomerulonephritis, with viral replication in mesangial cells and occasional focal necrosis of the renal tubular system; hyperplasia of the collecting ducts is associated with evident virus replication. Haemorrhages have been attributed to endothelial dysfunction, aggravated by virus replication in endothelial cells in the final stages of the disease. The renal interstitium displays intense oedema and an infiltrate largely composed of macrophages. Virus replication has also been observed in fibroblasts and in the smooth0muscle cells of arterioles and venules.In subacute-chronic forms of the disease, various types of glomerulonephritis are observed, ranging from mesangial proliferative glomerulonephritis to focal and segmental hyalinosis associated with immune-mediated phenomena. No striking changes are reported in the renal tubular system. Interstitial haemorrhages are associated with diapedesis due to immunologically mediated events. The interstitium has also been found to contain a lymphohistiocytic infiltrate with abundant plasma cells. No evidence has been reported of viral replication in any cell population.Abbreviations ASF African swine fever - ASFV African swine fever virus - C complement component - DNA deoxyribonucleic acid - HAD₅₀ 50% haemoadsorbing doses - Ig immunoglobulin - MPS mononuclear phagocytic system - PAS periodic acid-Schiff reagent  相似文献