| Avian leukosis of subgroup J is a highly contagious disease caused by a retrovirus,which first isolated from meat-type chicken. In recent years, a large number of reports showedthat infections of ALV-J were found in laying hens and local strains of chickens in ourcountry. In order to control and prevent ALV-J epidemic to protect the excellent local strainschicken, some purification measures need to be carried out in local chicken of China, butbecause the various strains and large population, non-standard and high density farming ofour country, it will take a long time to achieve very low infection rate in local chickens. Inorder to speed up the purification process, a variety of control methods and measures shouldbe taken out to reduce the rate of ALV-J infection. Vaccine prevention is an effectivealternative method, but so far, there has been no effective vaccine to prevent the ALV-J, sothis study aims to explore a useful vaccine in the use of ALV-J prevention.In this study, a pair of specific primers was designed based on the published ALV-J gp85gene sequence. The gp85gene was amplified by polymerase chain from the cDNA of Chinesestrain NX0101of subgroup J avian leukosis viruses, and cloned into the expressing plasmidvector pET32a (+). The recombinant plasmid vector was transformed into E. coli and IPTGwas used to induce the gp85gene expression. The expression production was purified andidentified by Western-blot with ALV-J specific monoclonal antibody JE9. The purifiedrecombinant protein with different adjuvants (F adjuvant and C adjuvant) was made intovarities of ALV-J subunit vaccines.7-day chickens and300-day hens were immuned withthese subunit vaccines and2-3weeks after the first immunization the second inoculation wascarried out. The serum antibody was tested weekly, after the second inoculation the chickenwas inoculated with the ALV-J, the fertilized eggs were collected and the hatched chickenwere inoculated with the ALV-J at1-day old. The viremia and the immunosuppressivedisease were detected to evaluate the immunopressive effection of the recombinant proteinsubunit vaccine; meanwhile the virus neutralization test of serum antibody was carried out invitro. The results are as follows:(1) ALV-J gp85gene which was amplified by polymerase chain reaction (PCR) is909bp,the recombinant protein expressed in E.coil DH5α was53KD, which can specific combinewith the ALV-J monoclonal antibody.(2) After inoculated to the recombinant protein vaccine with F adjuvant and F adjuvant, all the chickens could product the special serum antibody, the second inoculation couldenhance the titer of the antibody and prolong the antibody’s keeping time. The rate ofdecreasing viremia was53.8%and66.7%respectively after inoculated with the vaccines withF adjuvant and C adjuvant; those vaccines can also reduce the immunopathogenic of ALV-J.(3) The vaccine with F adjuvant and C adjuvant could induce the hens produce high titerserum antibody, the yolk antibody and the maternal antibody. The protections were65%and75%respectively in the hatched chickens infected the ALV-J in early with high titer maternalantibody, so the use of these vaccines can reduce the infection of ALV-J.The above results showed that the expressed recombinant protein combined differentadjuvants could reduce chickens and hens produce the neutralizing antibody, which canprovent ALV-J infection in early time. The study provided a scientific basis on the use ofsubunit vaccine in the breeding hens’ purification process. |
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