Construction of prokaryotic expression vector of canine distemper virus H gene

A vector containing canine distemper virus H gene (CDV H) was previously constructed in our lab and named pMD18-T-H. Based on the sequence of pMD18-T-H plasmid, a pair of primers with Xbal I and Hind III restriction site attached to the 5' end was designed and used to amplify the CDV H gene. The amplified fragment size of the CDV H gene was about 1800bp. The PCR product was purified and cloned into pMD18-T simple vector. Positive colonies were identified by using restriction endonuclease Xbal I and Hind III. Positive plasmid pMD18-Tsimple-H vectors was digested with Xbal and Hind III, and CDV H gene was recovered. At the same time, pPROEXTMHTa, pet-30b were digested with the same procedure. CDV H fragment was then ligated to pPROEXTMHTa and pet-30b respectively using T4 ligase. Positive colonies were again identified using Xbal and Hind III. Results indicated that the CDV H gene was cloned into two expression vectors successfully. This procedure lend experience for constructing expression vectors of CDV H gene in further studies, and would benefit for the production of CDV H protein for diagnosis and prevention of canine distemper.