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为了构建SmB/B’抗原表位的真核表达载体 ,通过体内、外进行表达并研究其免疫原性 ,采用异硫氰酸胍一步法提取BALB/c小鼠脾脏总RNA ,通过RT PCR方法克隆了含编码SmB/B’抗原表位的目的DNA ,构建了其真核表达质粒pcDNA3 fSmB/B’ ,体外转染HeLa细胞检测其表达及与抗体结合活性。体内采用基因免疫的方法对其免疫原性进行研究。结果 :成功构建pcDNA3 fSmB/B’真核表达质粒 ,Westernblot显示其在真核细胞内可有效表达且与抗SmB/B’抗体具有结合活性 ,经基因免疫实验组小鼠皆产生抗SmB/B’抗体。构建的SmB/B’抗原表位真核表达载体可在体内、外有效表达 ,且表达的抗原表位具有免疫原性
In order to construct a eukaryotic expression vector for SmB / B ’epitopes, the expression of SmB / B’ epitopes in E. coli was studied in vitro and in vivo. Total RNA was extracted from BALB / c mouse spleen by one-step guanidine isothiocyanate. The target DNA containing the SmB / B ’epitope was cloned and its eukaryotic expression plasmid pcDNA3 fSmB / B was constructed. The HeLa cells were transfected with HeLa cells to detect the expression of the SmB / B’ antigen and its binding activity to the antibody. In vivo immunogenicity of its method of immunogenicity. Results: The pcDNA3 fSmB / B ’eukaryotic expression plasmid was successfully constructed. Western blotting showed that it was efficiently expressed in eukaryotic cells and had binding activity with anti-SmB / B’ antibody. All the mice immunized with anti-SmB / B ’antibody. The constructed eukaryotic expression vector for SmB / B ’epitopes can be effectively expressed in vivo and in vitro, and the expressed epitopes are immunogenic