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目的获得人 Fas L蛋白 ,并对其功能进行初步研究。方法用 DNA重组法构建了 Fas L c DNA和谷胱甘肽转硫酶融合原核表达质粒 p GEX- KG- h FL。将重组质粒转入大肠杆菌 JM10 9,经 0 .2 mm ol/ L IPTG在 37°C条件下诱导 3h,SDS- PAGE检测。用 8m ol/ L尿素溶解的包涵体作为免疫原免疫家兔制备多抗。结果融合蛋白表达量约占细菌总蛋白的2 0 %。多抗效价达 1∶ 10 2 40 0 ,此多抗可以与重组蛋白发生很好的抗原抗体反应。结论 h FL 在大肠杆菌中得到高效表达 ,为深入研究 Fas L 提供了材料
Objective To obtain human Fas L protein and to study its function. Methods Fas LcDNA and glutathione S-transferase fusion prokaryotic expression plasmid pGEX-KG-h FL was constructed by DNA recombination. The recombinant plasmid was transformed into E. coli JM109, induced by 0. 2 mm ol / L IPTG at 37 ° C for 3h, SDS-PAGE. Polyclonal antibodies were prepared by immunization of rabbits with 8 mol / L urea dissolved inclusion bodies. Results The expression of fusion protein accounted for about 20% of total bacterial protein. Multi-anti-titer of 1:10 2 40 0, the polyclonal antigens and antibodies can react well with the antigen. Conclusion h FL was highly expressed in Escherichia coli, which provided material for further study of Fas L protein