尼罗罗非鱼TGF-β1蛋白原核表达及多克隆抗体的制备

Prokaryotic expression and polyclonal antibodies preparation of TGF-β1 protein in Nile tilapia

为制备尼罗罗非鱼TGF-β1多克隆抗体,采用同源重组技术构建尼罗罗非鱼pET-B2m-TGF-β1原核表达载体,转入大肠杆菌B21中诱导表达,将重组表达蛋白质纯化后免疫大耳兔制备多克隆抗体,采用Western Blot和ELISA检测抗体的特异性和效价。结果表明,构建的pET-B2m-TGF-β1原核表达载体经诱导表达获得了分子量为5.2×104的重组蛋白质,免疫大耳兔获得效价为1∶2048000的抗尼罗罗非鱼TGF-β1多克隆抗血清,该抗体能够特异性地识别原核表达的TGF-β1蛋白。

To prepare the polyclonal antibody against TGF-β1 of Nile tilapia,the TGF-β1 gene of Nile tilapia was transferred into the prokaryotic expression vector pET-B2m using homologous recombination technology,and transferred into Escherichia coli B21 for expression.The expressed TGF-β1 protein was purified and used to immunize rabbits for the preparation of polyclonal antibodies.The specificity and potency of the antibody were detected by Western blot and ELISA.The results showed that the pET-B2m-TGF-β1 prokaryotic expression vector was successfully constructed,and the recombinant protein with the molecular weight of 5.2×10^4 was obtained by inducing expression.The anti-Nile tilapia TGF-β1 polyclonal antiserum with a potency of 1∶2048000 was obtained from the immunized rabbit.The antibody could specifically recognize the prokaryotically expressed TGF-β1 protein.