Cloning and Expression of Tetanus Toxin Fragment C in E.coli.
The fragment C of tetanus toxin was amplified from Clostridium tetani DNA by PCR. This fragment was cloned into expression vector pET-28a(+),under the control of the T7 promoter. Expression of this plasmid in E.coli resulted in the production of a protein consisting of 6xHis of the vector fused to the N-terminal 451 amino acids of tetanus toxin. After induction with 1 mmol/L IPTG, TTC was expressed in E.coli BL21(DE3). The protein product accounted for 8.2% of the bacteria total protein in soluble form, SDS-PAGE and Western blot analysis of TTC recombinant protein confirmed this result. The expression products were also purified by Ni(2+)-IDA-Sephrose 6B column. Immunization of mice with rTTC resulted in the production of antibodies that were able to protect mice against a challenge with tetanus toxin furthermore, rTTC in vivo appeared to be able to undergo retrograde axonal transport.
He
Hua-Jun
HJ
Shanghai Institute of Biochemistry, the Chinese Academy of Sciences, Shanghai 200031, China. xfwu@sunm.shcnc.ac.cn
He
Zhi-Yong
ZY
Shi
Hui-Juan
HJ
Zhu
Wei
W
Yang
Guan-Zhen
GZ
Yuan
Qin-Sheng
QS
Wu
Xiang-Fu
XF