[Objective] This study aimed to investigate the outer membrane protein （OMP） patterns of Escherichia coli 038, 053 and 075 isolates from chickens. [Method] Eight pathogenic E. coil isolates with various serotypes we...[Objective] This study aimed to investigate the outer membrane protein （OMP） patterns of Escherichia coli 038, 053 and 075 isolates from chickens. [Method] Eight pathogenic E. coil isolates with various serotypes were used as experimental materials to extract OMP by using supersonic schizolysis method and Sarcosyl. After SDS-PAGE electrophoresis, OMP patterns of the extracted products were determined based on the OMP model diagram. [Result] OMP of eight E. coil isolates with three serotypes were divided into three patterns, to be specific, 2 075 isolates respectively belonged to OMP-I and OMP-II pattern, 1 053 isolate belonged to OMP-II pattern, and 5 038 isolates belonged to OMP-I and OMP-III pattern. [Conclusion] Experimental results showed that E. coli isolates with the same serotype may belong to completely different OMP patterns, while serologically unrelated isolates may belong to the same OMP pattern. OMP of E. coil isolates with the same serotype may generate genetic differentiation; in addition, OMP of E. coli isolates with different serotypes may have different genetic correlation.展开更多
Objective: To construct a recombinant plasmid containing the outer membrane protein 2 (Omp2) gene of Chlamydia trachomatis and express Omp2 in E.coli. Methods: The omp2 gene of C. trachomatis serovar D was cloned into...Objective: To construct a recombinant plasmid containing the outer membrane protein 2 (Omp2) gene of Chlamydia trachomatis and express Omp2 in E.coli. Methods: The omp2 gene of C. trachomatis serovar D was cloned into pQE30 vector following PCR amplification from genomic DNA. E. coli M15 transformants were induced to express the fusion protein by IPTG and the product was identified by SDS-PAGE and Western blot. Results: Confirmed by enzyme cleavage analysis and DNA sequencing, a correct recombinant plasmid pQE30/omp2 was constructed. The fusion protein from the transformants was approximately 60 kDa in size in SDS-PAGE analysis, which could specially react with anti-6 X His mouse monoclonal IgG antibodies. Conclusion: We successfully expressed Omp2 in E. coli M15, providing an efficient and simple system for assaying the immunological properties of Omp2.展开更多
以军团菌DNA为模板,PCR扩增获得军团菌主要外膜蛋白基因(M a jor ou ter m em brane prote in gene,om pS),与原核表达质粒pUC 18定向重组,构建重组质粒,转化大肠杆菌BL 21,并用限制性酶酶切分析、聚合酶链式反应、核酸序列分析、十二...以军团菌DNA为模板,PCR扩增获得军团菌主要外膜蛋白基因(M a jor ou ter m em brane prote in gene,om pS),与原核表达质粒pUC 18定向重组,构建重组质粒,转化大肠杆菌BL 21,并用限制性酶酶切分析、聚合酶链式反应、核酸序列分析、十二烷基磺酸钠-聚丙烯酰胺凝胶电泳、W estern印迹进行鉴定。实验结果表明我们扩增出了军团菌914 bp的om pS基因,成功构建了重组质粒pLPom pS,并在原核系统中得到了表达。展开更多
Objective: To directionally clone the ompl gene fromChlamydia trachomatis (Ct) F genotype onto a plasmid vectorfor constructing a rudimentary DNA vaccine. Methods: The complete ompl gene from genomic DNA of CtF genoty...Objective: To directionally clone the ompl gene fromChlamydia trachomatis (Ct) F genotype onto a plasmid vectorfor constructing a rudimentary DNA vaccine. Methods: The complete ompl gene from genomic DNA of CtF genotype wild species was amplified with primers designedby computer. The recombinant gene was obtained byrestriction enzyme cutting, linking the gene with the plasmidvector in vitro, transforming the recombinant gene intobacteria, and extracting the DNA from the bacteria. Results: DNA extracted from the bacteria was composed ofthe ompl gene and plasmid, which is identified by threemethods of singular restrictive enzyme cutting, doublerestrictive enzyme cutting and PCR. Conclusion: Cloning of the ompl gene from the Ct Fgenotype means that a rudimentary DNA vaccine wassuccessfully constructed.展开更多
基金Supported by China Postdoctoral Science Foundation(20100470565)Science and Technology Support Program of Hebei Province(10960408D)+1 种基金Project of Science and technology Bureau of Shijiazhuang(1150093A)Science and Technology Development Project of Qinhuangdao City(201101A182)~~
文摘[Objective] This study aimed to investigate the outer membrane protein （OMP） patterns of Escherichia coli 038, 053 and 075 isolates from chickens. [Method] Eight pathogenic E. coil isolates with various serotypes were used as experimental materials to extract OMP by using supersonic schizolysis method and Sarcosyl. After SDS-PAGE electrophoresis, OMP patterns of the extracted products were determined based on the OMP model diagram. [Result] OMP of eight E. coil isolates with three serotypes were divided into three patterns, to be specific, 2 075 isolates respectively belonged to OMP-I and OMP-II pattern, 1 053 isolate belonged to OMP-II pattern, and 5 038 isolates belonged to OMP-I and OMP-III pattern. [Conclusion] Experimental results showed that E. coli isolates with the same serotype may belong to completely different OMP patterns, while serologically unrelated isolates may belong to the same OMP pattern. OMP of E. coil isolates with the same serotype may generate genetic differentiation; in addition, OMP of E. coli isolates with different serotypes may have different genetic correlation.
基金This work was supported in part by grants from the Department of Science and Technology of Hunan Province (No. 01SSY2008-6) the Department of Health of Hunan Province (No. B2003-078).
文摘Objective: To construct a recombinant plasmid containing the outer membrane protein 2 (Omp2) gene of Chlamydia trachomatis and express Omp2 in E.coli. Methods: The omp2 gene of C. trachomatis serovar D was cloned into pQE30 vector following PCR amplification from genomic DNA. E. coli M15 transformants were induced to express the fusion protein by IPTG and the product was identified by SDS-PAGE and Western blot. Results: Confirmed by enzyme cleavage analysis and DNA sequencing, a correct recombinant plasmid pQE30/omp2 was constructed. The fusion protein from the transformants was approximately 60 kDa in size in SDS-PAGE analysis, which could specially react with anti-6 X His mouse monoclonal IgG antibodies. Conclusion: We successfully expressed Omp2 in E. coli M15, providing an efficient and simple system for assaying the immunological properties of Omp2.
文摘以军团菌DNA为模板,PCR扩增获得军团菌主要外膜蛋白基因(M a jor ou ter m em brane prote in gene,om pS),与原核表达质粒pUC 18定向重组,构建重组质粒,转化大肠杆菌BL 21,并用限制性酶酶切分析、聚合酶链式反应、核酸序列分析、十二烷基磺酸钠-聚丙烯酰胺凝胶电泳、W estern印迹进行鉴定。实验结果表明我们扩增出了军团菌914 bp的om pS基因,成功构建了重组质粒pLPom pS,并在原核系统中得到了表达。
文摘Objective: To directionally clone the ompl gene fromChlamydia trachomatis (Ct) F genotype onto a plasmid vectorfor constructing a rudimentary DNA vaccine. Methods: The complete ompl gene from genomic DNA of CtF genotype wild species was amplified with primers designedby computer. The recombinant gene was obtained byrestriction enzyme cutting, linking the gene with the plasmidvector in vitro, transforming the recombinant gene intobacteria, and extracting the DNA from the bacteria. Results: DNA extracted from the bacteria was composed ofthe ompl gene and plasmid, which is identified by threemethods of singular restrictive enzyme cutting, doublerestrictive enzyme cutting and PCR. Conclusion: Cloning of the ompl gene from the Ct Fgenotype means that a rudimentary DNA vaccine wassuccessfully constructed.
