Cloning of adult rat Nogo gene segments and expression and purification of their coding protein fused with GST

    ZHANG Ping, CHENG Xiª²Ping, FEI Lingª²Ling, WANG Chunª²Ting, YANG Hao, CHEN Binª²Fu, JU Gong

    Institute of Neuroscience, School of Basic Medicine, Fourth Military Medical University, Xi¡¯an 710033, China

    ¡¾Abstract¡¿ AIM: To obtain rat Nogo gene segments, express efficiently in E. coli and purify the target proteins. METHODS: Nogo gene segments were amplified by RTª²PCR from normal adult SD rat spinal cord total RNA. After sequenced, the reconstructed expression vectors were constructed by enzyme digestion and subcloned into expression vector pGEXª²4Tª²1. Then the vectors were transformed into E. coli DH5¦Á. Recombinant GST fusion proteins were expressed via the induction of IPTG and purified through glutathione agarose column. RESULTS: The sequences of cloned adult rat Nogo gene segments (570-691 and 1028-1089 amine acid residues) were identical with those earlier reported. Two protein bands of Mr39 000 and 32 000 appeared on SDSª²PAGE gel after the expressed GST fusion proteins were separated by SDSª²PAGE. Then the expressed fusion proteins were purified to high purity. CONCLUSION: The adult rat Nogo gene segments have been successfully cloned and efficiently expressed in E. coli, and the GST fused target proteins have been purified to high purity. ......