| Customarily, the prevention measures to AL are supposed to approach from three basic aspect—the infection sources,the dissemination path, susceptible animal. But considering the basic situation of our country’s aquaculture industry, it is hard to control the sources as well as curing the animal, and cutting the path is a kind of potential and effective method. For the moment, no researches or preptors regarding RNA intereference to virus receptors have been observed nor found by the author in the available bibliography, and we have not found the positive function after consulting to many reference. Accordingly we designed si RNA which restrain TVBreceptor peculiarly, and consequently prevent the infection of ALV-B.The viral Env of ALV-B interacting with TVB receptor are linked to activation of death receptor signaling pathways on cell membrane and induced to apoptosis, considering this character, we used RNA interference technique to decrease the m RNA specifically which transcribe to TVB receptor and lead to the consequence that the cell reduce to express TVB receptor which medicate ALV-B binds.Two models of TVB receptor can medicate ALV-B binds in the body of chicken, called TVBS1 and TVBS3, Achieving the aim that against virus infection, we designed two small interference RNA according to the m RNA respectively which transcribe to TVBS1 and TVBS3. Then process them to short hairpin RNA according to experiment requirements. Link sh RNA to the p Silencer 2.1-U6 plasmid and named them Tp S1-1, Tp S1-2,Tp S3-1,Tp S3-2 respectively, composed a control plasmid randomly. We set the recombinant plasmids in different group following the experiment plan, transfected into DF-1 cell then get the cell line which can produce recombinant plasmids stably after 10 day’s preparation by screening. Indirect immunofluorescence was used to detect the situation of virus copy after 7 days challenge of ALV-B. We set three groups that inoculated positive plasmids and one group inoculated negative plasmid for animal experiment. Chickens were inoculated in accordance the plan cut the age of 1 day and 4 days. The virus ALV-B was inoculated at the age of 7 days, then made autopsy every 6 days and inspected 5 times eventually. Exploring the effect that RNAi against copy of ALV-B by consecutively testing and analyzing to body weight, immune organ index, antigen of p27, CD4+ and CD8+ lymphocyte.We observed the amount of fluoresecence decreased in the group 1 compared to the control group, which were no significant difference between group 2, group 3 and control group. This experiment part preliminary prove that group Tp S1-1 and Tp S3-1 inhibited the synthesizing of TVB receptor, group Tp S1-1 and Tp S3-2, group Tp S1-1 have little significant effect to it. The experiment result shows that chickens in group 1 have certain resistance to ALV-B, which gives indirect evidence that the combined action of Tp S1-1 and Tp S3-1 produce positive interference effect to TVB. While group 2 and group 3 are liteele different from the control group, indicate that the combined action Tp S1-1 and Tp S3-2, the single action of Tp S1-1 have insignificant interference effect to TVB receptor.Eventually come to the conclusion, the combind action of Tp S1-1 and Tp S3-1 can effectively inhibit the production of TVB in the cells as well as body, while the effect of inhibition engendered by any single plasmid is not markedly. |
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