Isolation And Identification Of Porcine Pseudorabies Virus Strain YuA And Cloning And Sequence Analysis Of The PK Gene From Strain YuA

A strain of virus was isolated from a pig farm in Henan.The isolate was continually inoculated 4 generations onto PK15 cell and induced typical cytopathogenic effect(CPE).Three rabbits were injected with the isolates and no clinical sign was observed.The titers of the antibody to Prv in rabbits were 1:4,1:4 and 1:16 by LA 20 days post-challenge. The DNA extracted from the isolated virus was subjected to PCR amplifying ,the right fragment was amplified.All of the evidence indicated that the isolated virus was pseudorabies virus,and it was named strain YuA of pseudorabies virus. A pair of primer were synthesized according the PK gene sequence of strains Er and NIA-3 published on GeneBank.PK genes of Prv strains MinA,YuA and YuB were amplified and cloned into vector pMD 18-T .These rocombinant plasmids were identified by restriction enzyme analysis and sequencing. By means of molecular biology software,the PK genes of 3 Prv strains YU-A,Yu-B and Min-A were compared with that of another 3 strains(Er,NIA-3 and Ka) of Prv and were multialigmented.The results indicated that the identities between the 6 strain was high.The lowest identities (97.1% )was between strains YuA and Ka .A deletion of base G at the site 282th in Prv strain YuA accounted the evident difference in the amino acid sequence from the 93th Aa of YuA strain from the other strains.The deduced amino acid identities of strain YuA with strains MinA,YuB,Er,NIA-3 and Ka were 30.7%,30.7%,30.2%,29.5%and19.5%,respectively, much less than the lowest identity (86.1% )between the other strains.The mutant of PK gene of strain YuA may have some relation with the rabbits' showing no clinical sign.The nucleotide sequence analysis of the PK gene of the Prv strains revealed that there are 2 highly variable regions which lie on 238-249 nt and 1081-1098 nt in their nucleotide sequences.