中国兽医科学2025，Vol.55Issue(4)：492-499,8.DOI:10.16656/j.issn.1673-4696.2025.0066
多杀性巴氏杆菌OmpH蛋白间接ELISA方法的建立
Establishment of an indirect ELISA method for Pasteurella multocida based on OmpH protein
摘要
Abstract
This experiment induced the OmpH antigen of Pasteurella multocida in the prokaryotic expression system of Escherichia coli,and established an ELISA detection method for bovine Pasteurella multocida antibodies,providing strong scientific basis for the development of immunological antibody detection methods for Pasteurella multocida.We analyzed the signal peptide and transmembrane hydrophobic region sequence of OmpH sequence and performed rare codon optimization on the selected target gene fragment.The chemically synthesized nucleotide sequence was connected to the pET32a vector,and constructed the recombinant plasmid pET32a-OmpH.The recombinant plasmid was transformed into the competent cell line of Escherichia coli BL21(DE3),and expanded cultivation and successfully induced the expression of high purity inclusion body OmpH protein with a size of about 57 kDa and a protein purity of about 94％.The OmpH recombinant protein has no cross reactivity with other disease antibodies.The indirect ELISA detection method established using the purified pET32A OmpH recombinant protein as the coating antigen has strong specificity and no cross reactivity with other bovine pathogens.The intra-and inter group coefficients of variation are both below 10％,which means the method indicating good stability and reproducibility.This study lays the foundation for further development of mature multi killing Pasteurella antibody detection kits.关键词
多杀性巴氏杆菌/OmpH蛋白/原核表达/蛋白纯化/ELISAKey words
Pasteurella multocida/protein OmpH/prokaryotic expression/protein purification/ELISA分类
农业科技引用本文复制引用
王润清,孙航,阚威,李歌,李琦,侯晓林,孙雨..多杀性巴氏杆菌OmpH蛋白间接ELISA方法的建立[J].中国兽医科学,2025,55(4):492-499,8.基金项目
国家现代农业产业技术体系建设专项(CARS39) （CARS39）
