Construction Of Anti-canine Distemper Virus Single Domain Antibody Library And Prokaryotic Expression Plasmid Of Canine Distemper Virus Gene

Canine distemper is a highly contagious viral disease caused by canine distemper virus, which belongs to measles virus in paramyxoviridae. Its main clinical symptoms include diphasic fever, erythema, conjunctivitis, leukopenia and central nervus system damage. In recent years, the effect of canine distemper vaccine is decline gradually, and the monoclonal antibody is widely used since its high specificity in the diagnosis and treatment. So, it is great significance to research and develop the canine distemper diagnositic and therapeutic reagents.Heavy chain antibody that naturally lack of light chain was existed in camel, and the advantages make them has the extensive antigen binding ability. Phage display antibody library technology is one of the most outstanding achievements in the field of antibody engineering in recent years. The technology mainly display antibody on the surface of phage, and screen the specific antibody with antigen,and followed with expression and identification the antibody function, so make the gene engineered antibody preparation more convenient and quick.The study aims were to prepare CDV specific single-domain antibodies for developing CDV detection method, and passive immunization reagent with its neutralizing activity to prevent canine distemper. In this study, we used canine distemper virus to vaccinate bactrian camel, and then separated the lymphocytes from the peripheral blood and extracted total RNA, then synthesized cDNA through RT-PCR, then used single-domain antibody specific primer to obtain VHH gene fragment after three times amplification. VHH gene was inserted into the pCANTAB5E phagemid, and the recombinant plasmid was electro-transformed into E.coli TG1. W constructed a canine distemper VHH antibody library with a library size of 1.3×106. We also cloned the canine distemper ’virus key antigens H、F、N to prokaryotic expression vector pET28a, and recombinant plasmid was transformed into competent cell BL21 (DE3), and the constructed expression plasmid will provide a good foundation for the antibody screening from the library, and which providing the new idea for the treatment of canine distemper.