Involvement Of Molecular Adjuvants In Somatostatin Gene Immunization

With constructing the fusion expression plasmid harboring GM-CSF and S/2SS based on the fusion expression plasmid harboring somatostatin (SS) and HBsAg S( pES/2SS ), gene adjuvant vectors and preparation of the E.coli DNA and so on, experimental mice and Hu Lambs were immunized in the current study to investigate the factors influencing SS gene immune response and its effects on the body growth and endocrine states. An appropriate immune protocol was developed to promote animal growth and accelerate clinical application for economical, effective and safe methods of SS gene vaccines.1. Construction and identification of fusion expression plasmids harboring GM-CSF and S/2SS and gene adjuvant plasmidsâ‘ Prediction of antigenic epitope on the fusion proteinThe secondary structure and physical and chemical characteristics of the expected fusion protein 2SS-HbsAg(S) in pES/2SS-GMCSF and pES/2SS plasmids were analyzed by DNA Star software to predict epitopes. Both of them are the same, the epitopes of fusion protein 2SS/S were probably at the amino acid serial where SS were inserted regarding its hydrophobicity, antigenicity and hydrophilicity etc. Besides fusion protein 2SS-HBsAgS, the other epitope at amino acid serial possessed the fusion protein S/2SS-GMCSF. So, the GMCSF may not change epitopes of 2SS/S fusion protein and enhance the immune effect on SS gene vaccine. In addition, CpG DNA and other adjuvant can enhance the immune effect of SS DNA vaccine in theory.â‘¡Construction and identification of fused expression plasmid of pES/2SS-GMCSF and gene adjuvant plasmidsGMCSF gene was cut by enzymes of Xho I and EcoR I from pcS/2SS-GMCSF, and was directly inserted into vectors of pES/2SS (terminal condons between S/2SS and GFP) and pEGS/2SS(no terminal condons and GFP could be expressed), the recombinant pES/2SS-GMCSF and pEGS/2SS-GMCSF were identified by restriction endonuclease digestion and sequencing. In a same way, GMCSF gene was cut by enzymes of Xho I and EcoR I from pcGMCSF, and CpG gene with enzyme site sequence of EcoR I and Sal I was synthesized chemically and turned double chain annealed. Subsequently, both of them were inserted into the pEGFP-Nl, respectivelyand were constructed the recombinant adjuvant plasmids of pE-GMCSF and pE-CpG. The insertion site, orientation and sequence were comformed by restriction endonuclease digestion and sequencing. These recombinant...