抗BAFF单克隆抗体轻链和重链可变区基因的克隆及鉴定

Cloning and identification of the variable region gene of mouse anti-human BAFF mAb

目的:在本实验室成功制备小鼠抗人BAFF单克隆抗体的基础上,通过分子克隆方法获得该抗体可变区基因序列。方法:从1株小鼠抗人BAFF单克隆抗体杂交瘤细胞FMMUB4中提取总RNA,以此为模版反转录成cDNA,用针对小鼠单克隆抗体重链和轻链可变区基因序列的特异性引物分别进行PCR反应,PCR产物连接入载体,经筛选阳性克隆、酶切鉴定后送测序,测序结果进行生物信息学分析。结果:成功克隆了抗BAFF单克隆抗体FMMUB4重链和轻链可变区基因,测序结果证实序列正确。结论:获得了抗BAFF单克隆抗体FMMUB4重链和轻链可变区基因序列,为下一步构建相关基因工程抗体打下良好基础。

AIM： To obtain the variable region gene sequence of heavy and light chain of mouse anti-human BAFF monoclonal antibody （mAb） on base of BAFF mAb which was cloned in our laboratory. METHODS： The total RNA was extracted from mouse anti-human BAFF mAb hybridoma cell line FMMUB4, and then the RNA was reverse tran- scribed into cDNA. Specific primers were designed to amplify the targeted gene. The targeted gene fragments were inserted into vectors to construct the clone vectors. The gene sequences were analyzed after identified by positive clones screening and restrictive enzyme digestion. RESULTS： The variable region gene sequences of mouse anti-human BAFF mAb were obtained. CONCLUSION： The variable region gene sequences of mouse anti-human BAFF mAb will provide experimental basis for further study on constructing engineered antibodies.