ZHAO Bosheng, ZHANG Shicui, PANG Qiuxiang, LIU Zhenhui, LIANG Yujun. Expression, purification and polyclonal antibody generation of p23, an Hsp90 cochaperone, in the amphioxus Branchiostoma belcheri[J]. Acta Oceanologica Sinica, 2006, (6): 99-105.
Citation:
ZHAO Bosheng, ZHANG Shicui, PANG Qiuxiang, LIU Zhenhui, LIANG Yujun. Expression, purification and polyclonal antibody generation of p23, an Hsp90 cochaperone, in the amphioxus Branchiostoma belcheri[J]. Acta Oceanologica Sinica, 2006, (6): 99-105.
ZHAO Bosheng, ZHANG Shicui, PANG Qiuxiang, LIU Zhenhui, LIANG Yujun. Expression, purification and polyclonal antibody generation of p23, an Hsp90 cochaperone, in the amphioxus Branchiostoma belcheri[J]. Acta Oceanologica Sinica, 2006, (6): 99-105.
Citation:
ZHAO Bosheng, ZHANG Shicui, PANG Qiuxiang, LIU Zhenhui, LIANG Yujun. Expression, purification and polyclonal antibody generation of p23, an Hsp90 cochaperone, in the amphioxus Branchiostoma belcheri[J]. Acta Oceanologica Sinica, 2006, (6): 99-105.
Department of Marine Biology, Ocean University of China, Qingdao 266003, China;School of Life Sciences, Shandong University of Technology, Zibo 255049, China
2.
Department of Marine Biology, Ocean University of China, Qingdao 266003, China
The cDNA of amphioxus p23, a highly conserved co-chaperone for Hsp90, was cloned into a bacterial expression vector pGEX-6P-1 and the GST-tagged fusion protein was produced in Eschherichia coli cells.The recombinant p23 was purified by affinity purification, and its molecular mass was estimated to be approximately 22 kDa by sodium dodecyl sulfate-polyacrylamide gel electrophoresis.The N-terminus of purified p23 was sequenced, and the resulting amino acid sequence matches exactly the predicted residues deduced from the amphioxus p23 gene.Besides, polyclonal antibodies against the recombinant p23 were generated, and these antibodies not only recognized specifically the fusion protein GST-p23 from induced E.coli cells, purified GST-p23 and p23 protein, but also reacted with the total protein extracted from the adult amphioxus and formed a single positive band.These results pave the way for identifying its tissue and subcellular localization, and may open the door to clarifying its structure and mechanisms of biological role.
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