顶头孢霉cefG基因的表达及抗体制备的研究

Recombinant expression and antibody preparation of cefG by Acremonium chrysogenum

通过PCR方法得到带酶切位点的cefG基因,用于构建表达质粒pET30-G,并在大肠杆菌里成功表达出乙酰转移酶的包涵体。将该重组蛋白的标准品作为抗原,免疫BALB/c小鼠得到多克隆抗体,酶联免疫吸附测定(ELISA)分析得抗体的效价满足实验需求。利用该抗体绘制出抗原浓度的标准曲线,确定待测抗原适合的检测浓度在0.16μg/mL以下。制得的抗体在重组大肠杆菌里检测到了乙酰转移酶,证明所得抗体具有实验价值。

Amplification by the polymerase chain reaction（PCR） of the cefG gene with a restriction enzyme cutting site has been used to construct the expression plasmid pET30-G,which expressed acetyltransferase in E.coli in the form of inclusion bodies.The recombinant protein was then vaccinated into BALB/c mice to obtain polyclonal antibodies.Satisfactory antibody titers were obtained by ELISA analysis.The calibration curve of the antigen concentration was drawn using this antibody,and indicated that the optimum concentration of the antigen is below 0.16μg/mL.Acetyltransferase was detected in the E.coli recombinants,which indicates that the antibody is useful for practical studies.