Effect On Immunization Of Mice Or Cattle Against Inactivated E.coli (dh5a) Carrying Inhibin Plasmid Pcis

In this study, Techniques such as gene immunization, bacterial carriers, enzyme-linked immunoassay (ELISA) and B-type ultrasonograph were used to study the effect of immunization against inactivated E. coli DH5a carrying inhibin plasmid pCIS on immune response,reproduction performance and gene safety. By it, we try to find the optimal strategy of inhibin gene immunization and then set a base for improving the reproductivity of animal. The main content presents as follows:1, Study on immunization of mice against inactivated E.coli (DH5a) carrying inhibin plasmid pCISHigh doses of plasmid DNA were necessary to induce immune response in large animals. Because production of DNA vaccines is a tedious process, so a new strategy were found that inhibin gene vaccines were delivered by inactivated E.coli DH5a. In this study, mice were divided into five groups including three experimental groups and two control groups. There were three different doses in every experimental group. The experimental groups were injected pCIS/PC, pCIS/DDA and inactivated bacteria respectively. The results showed that the P/N value of mice immunized with E. coli DH5a-mediated plasmid reached 2 after the primary immunization. There were no significant difference among three different doses (P＞0.05). The P/N value of mice vaccinated with 50μg of pCIS/PC was 2.435±0.309 which was higher than other two doses (P＜0.05). The effects were the same in inhibin plasmid with DDA. However, no significant differences were observed between the best doses of plasmid group and the inactivated groups (P＞0.05). All the vaccinated groups were significantly different in antibody P/N value from the control groups (P＜0.05). Simultaneously, there were no significantly different in antibody P/N value between the primary immunization and the secondary immunization (P＞0.05). with respect to percent of Mice with positive antibody against inhibin, there was no significant difference in inactivated group (P＞0.05). However, there was significant difference in different dose of the plasmid groups (P＞0.05). After the secondary immunization, the maximum percent achieved 90% and the lowest was 50% in pCIS/PC group, however, 30% in pCIS/DDA group. In Mouse litter size, it is higher in the inactivated groups than the control and the plasmid groups but no significant difference (P＞0.05). The best group is 50μg of pCIS/PC group. About safety, comparing the experimental groups with control, there were no difference in heart, liver, spleen, lung and kidney (P＞0.05) suggesting the new strategy was safe.In summary, though the effect had no significant difference between the plasmid groups and the inactivated groups, the dose of injection is very low in activated group suggesting the new strategy was feasible that mice were vaccinated with E.coli DH5a-mediated plasmid.2, Study on immunization of cattle against inactivated E.coli (DH5a) carrying inhibin plasmid pCISBecause E.coli DH5a-mediated DNA vaccine was able to induce strong immune response in mice, in this research, we detected the feasibility of this method in cattle by comparing with the group of PMSG and DNA vaccine with DDA. The results reveal the P/N value was all low after the first immunization, had an obvious increase after the second immunization and then decrease in the following days. In the groups of inactivated bacteria, the P/N value had no significant difference among the doses (P＞0.05), no significant difference comparing with the plasmid groups (P＞0.05). The number of positive cattle was 2 in pCIS/PC and pCIS/DDA groups after the first immunization, and 3 after the second immunization. In inactivated groups, the number of positive cattle was the best in the high dose group. The ovaries were detected by B-type ultrasonograph in oestrus cycle of cattle. The number of mature follicles is more in PMSG group. However, no remarkable difference between groups of inactivated vaccines and groups of plasmid DNA (P＞0.05). In PMSG group, rate of two follicles was 60% (6/10), but 20% in control. In the low dose of inactivated groups, rate of two follicles reached 30% (3/10). The rate of two follicles was increased following the dose. There were no remarkable difference concerning numbers of mature follicles between the middle dose of inactivated vaccines and the control (P＞0.05), but had a significant difference in other dose groups (P＜0.05).In summary, although significant difference were observed between groups of E. coli DH5a-mediated DNA plasmid and PMSG, no significant difference compared with plasmid with adjuvants (P＞0.05). Because the immune effect induced with the highest dose of inactivated vaccines was better than the other in this study, so we didn't confirm which is the optimal immune dose. Therefore, the ability of inhibin plasmid pCIS using for E. coli DH5a as carrier to stimulate strong humoral and cellular immune responses warrants further studies.