Preparation And Detection Of Dendritic Cell-targeting Peptides Recombinant NDV HN Gene Lactobacillus

Newcastle disease (ND),which is found worldwide and causes one of the most prevalentinfectious diseases，it is also more common in our country and caused significant economic lossfor breeding industry.In this study, we selected a foreign protein expression vector—Lactobacillus (Lactobacillusplantarum NC8) to deliver the gene of Newcastle disease virus HN as major protective antigenprotein integrates with Dendritic cell-targeting peptides (DCpep), combined with exogenousgene specific immune function and probiotic lactic acid bacteria developing recombinantlactobacillus oral vaccine of DCpep fusion HN gene, and preliminary analysis of the immunefunction.By means of primer5.0software design primers in this experiment, we using RT-PCRamplified gene fragment of Lasota strain HN, connected with the shuttle expression carrierpSIP409after the fusion of the target gene and36bp of Dendritic cell-targeting peptides(DCpep) by enzymes slash,PCR and sequencing analysis to identify, the target recombinantplasmid electroporated into Lactobacillus plantarum, after analysising by SDS-PAGE, Westernblotting and flow cytometry to further detect exogenous gene, two fusion proteins ofpSIP409-HN and pSIP409-HN-DC presented reactionogenicity as the polyclonal antibody ofNewcastle disease virus, which obtained the target protein by SppIP peptide inducing expressionin lactic acid bacteria, indicating that it has laid the foundation of constructing recombinantNDV lactobacillus oral preparation.To further analyze the immune protective effect of recombinant lactic acid bacteria, the9-day-old chicks were randomly divided into four groups, according to experimental designprocedures were respectively immunized with pSIP409-HN-DC, pSIP409-HN, pSIP409andPBS. By detecting the level of HI titers, contents of sIgA in intestine and trachea, eachsub-category changes in spleen and peripheral blood CD3+and CD4+T cells and T lymphocytesproliferation in vitro after the body’s immune, indicating that the recombinant bacteria immuneeffect of DCpep fusion HN gene is better than mere HN.After challenge by dynamicallymonitoring the mental state of the chickens, weight levels, survival and histopathologicalchanges, etc. for immune parameters of each experimental group, we conducted a more objective analysis. Statistical results show that the immune protection factors ofpSIP409-HN-DC group,better than the pSIP409-HN group, and the two experimental groupswere higher than other control groups, indicating that DCpep as a vaccine adjuvant has theeffect of enhancing immune function,also showed that recombinant genetic engineering oflactic acid bacteria could made chickens obtain a certain degree of ability to resist NDV attack,the research provided dara for further study of enhanced immune ND genetically engineeredLactobacillus oral vaccine.