重症肌无力单链抗体基因的制备

Preparation of single chain variable fragment gene in myasthenia gravis

[目的 ]构建抗人乙酰胆碱受体主要免疫原区单链抗体 .[方法 ]应用多聚酶链反应从抗人乙酰胆碱受体主要免疫原区抗原结合片段扩增重链和轻链可变区基因 ,并克隆到载体噬粒 pHEN2 .将含有单链抗体基因的重组噬粒转化大肠杆菌DH5α ,分离提纯重组噬粒后再经酶切、琼脂糖凝胶电泳检查以确认单链抗体基因的正确性 .[结果 ]电泳检查发现了大小分别为 3 78bp ,3 3 9bp和 762bp的重链可变区、轻链可变区和单链抗体基因 ,且位置正确 .[结论

OBJECTIVE?To construct the gene of a single chain variable fragment against the main immunogenic region in acetylcholine receptor.?METHODS?The heavy chain and light chain variable region genes were amplified from an human antigen binding fragment against the main immunogenic region in human acetylcholine receptor using PCR, and cloned in vector phagemid pHEN2. The recombinant phagemid containing the single chain variable fragment gene was subsequently transformed into E coli DH5α. After isolation and purification, the recombinant was digested with endonucleases and checked in agarose gel electrophoresis to confirm the right size of the single chain variable fragment.?RESULTS?The sizes of the heavy chain variable region gene, the light chain variable region gene and the single chain variable fragment gene were found to be approximately 378?bp, 339?bp and 762?bp respectively, and the inserted positions were correct.?CONCLUSION?A single chain variable fragment gene against the main immunogenic region in human acetylcholine receptor has been successfully constructed .