Establishment Of Rapid Detection Methods, Cloning And Sequencing Of F Gene For Canine Distemper Virus

CD(canine distemper) was a kind of highly contagious disease caused by cdv(canine distemper virus) infection, which was one of the most harmful loemias for Pets,Fur Animal and Conservation of Wildlife. Because there were many difficulties for viral isolation and clinical final diagnosis of CD,this experiment established molecular biology detection methods for cdv(canine distemper virus) to solve the problem of nosogenic diagnosis.The focal point was to establish detection methods of RT-PCR(reverse transcription polymerase chain reaction) and FQ-PCR (Flurogenic Quantitative Polymerase Chain Reaction)to provid technical insurance for human health as well as pets and other animals well growth.We collected many kinds of nosogenic canine excreta such as blood,urine,saliva ,nasal swab,eye secretions and so on from pet clinic in Fujian areas.We carried out detecting canine distemper virus by RT-PCR,SYBR Green I RT-PCR,FQ- PCR methods that was established through designing primers on the basis of published F gene sequences and the results accorded with the theory afterelectrophoresis. These three methods had high specificity and saitivity so that they could be applied on the clinical rapid detection of CDV(canine distemper virus).In addition, three pairs primers were designed to amplify F gene complete sequence from wild strains.The depurated reclaiming products were transformed to E.coli DH5αthrough PMD18—T vector which has contained sequences by clone.After that,we analyzed the variation of F gene 's conservative sequence by metered results of Shanghai bioengineering company and revealed the gene difference among strains.All the conclusions have important theory and practical value for the identification of CDV,epidemiological investigation,development of vaccine and research of clinical diagnosis.