The Study On Immune-Regulating DNA Vaccine PcDNA4.0-MZ-TGF-β₄ Against Eimeria Tenella

The coccocidiosis of chicken is a cytoplastic entozoic parasite disease and may cause severe economic loss to the industry of poultry . Current methods for coccidiosis control in the poultry industry have many drawbacks, such as drug-resistance and the potential reversion to virulence. Therefore, DNA vaccines containing important parasitic antigens have been suggested as an alternative to the virulent or attenuated vaccines. Chicken TGF-β₄ is an important cytokine which has function on renovation and promotion after damnification. The MZ5-7 is one of the antigens of Eimeria tenella second generation merozoites, and has been shown to elicit partial protection in chicken against an oral challenge with E.tenella, In this report, we constructed immune regulative DNA vaccine by the combination of Chicken TGF-β₄ gene with MZ5-7 gene of E.tenella and tested their protective effects on the chicken against the challenge of E.tenella.Using the spleen lymphocytes (SP) total RNA of chickens infected firstly with E. tenella sporulated oocysts (about 5×10⁴ per chicken) as the template, the gene of chicken transforming factor- β was amplified and cloned through the RT-PCR. The PCR products were then cloned into pMD18-T vector for sequencing. The gene ORF was 345 bp, and it matched with the predicted size. Compared with the published sequence in GenBank (M31160), the homology of nucleic acids and amino acids were 99.7% and 100% respectively. The gene was later sub-cloned into a prokaryotic expression vector, designed as pET28a-TGF-β₄. After induction by IPTG , the ChTGF-β₄ gene was expressed and the fused protein was 17 kD.By the means of DNA recombination technique, pcDNA4.0-MZ-TGF-β₄ and pVAX1.0-TGF-β₄ vectors were constructed. After identified by restriction enzymes...