The invention discloses a
canine distemper virus CDV-3 strain infectious
cDNA cloning and constructing method and application thereof. A
canine distemper virus CDV-3 strain is used as a template, theoverall length of the CDV-3 is divided into 5 fragments, and RT-PCR amplification is performed. Through
enzyme-
cutting splicing, the 5 fragments are sequentially inserted into a eukaryotic vector, besides, a hammerhead
ribozyme sequence and a
hepatitis D
ribozyme sequence are respectively added to an F1 head end and an F5
tail end, and
canine distemper virus CDV-3 strain infectious cDNA clone is obtained; then three auxiliary plasmids expressing CDV-3N, P and L proteins are constructed; the
canine distemper virus CDV-3 infectious cDNA clone and the three auxiliary plasmids are used for performing cotransfection on 293T cells so that saved recombinant CDV-3 viruses (rCDV-3) are obtained. Through research, the highest
titer of the obtained rCDV-3 virus can reach 107.667TCID50 / mL, which is 10times higher than that of wtCDV-3. After Vero cells are infected with the rCDV-3, within 36h after infection, the highest virus
titer can be achieved; but after the Vero cells are infected with the wtCDV-3, within 72h after infection, the virus content can reach the highest value. A base is established for developing a canine distemper virus novel vaccine and studying the infective mechanism through the
canine distemper virus CDV-3 strain infectious
cDNA cloning and constructing method disclosed by the invention.