Expression of Duck dTLR2 in Immune Organs and Blood and Preparation of Polyclonal Antibodies
In order to analyze the possible role of dTLR2 in duck innate immunity,we study the expression of dTLR2 mRNA in immune organs during 1 to 10 weeks,as well as the expression in blood after challenging with Escherichia coli and Salmonella enteritis by Fluorescence Quantitative PCR.We also prepared polyclonal antibodies of the extracellular domain of dTLR2 by recombinant protein expression for further studying the biological function of dTLR2.The results showed that dTLR2 mRNA was expressed in spleen,thymus and bursa at different weeks.The expression of dTLR2 mRNA was significantly different among different weeks in the spleen and thymus tissues,and there was no significant difference in bursa.After infection with E.coli and S.enteritis,the expression of dTLR2 mR-NA was significantly higher than that of the control group in the first and second days,but there was no significant difference among groups in the third day.Analyzing the sequence of dTLR2 gene,we predicted the extracellular do-main,designed primers for PCR assay,and built pET28a-TLR2 recombinant plasmid.The recombinant plasmid was translated into E.coli RL21(DE3)for expression of dTLR2-His protein.The SDS-PAGE results showed that the fu-sion recombinant protein efficiently expressed in the supernatant,molecular mass was around 29 ku.The results of Western Blot showed that the antibody which harvested from immunizing rabbits could detect recombinant dTLR2 and endogenous dTLR2.The analysis of the expression of dTLR2 and successful preparation of polyclonal antibody provide favorable support for further studying of dTLR2 biology function.
万方数据
维普
