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《Chinese Journal of Biotechnology》 2005-02
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Expression and Bioactive Characterization of Bacteriophage Lysin Gene of Bacil lus anthracis in Escherichia coli

LI Xiao-Jing 1,2, ZHANG Hao1, FU Xue-Qi2, LI Yan-Ying1, CHEN J ing1, LI Yu-Ling1, FANG Hong-Qing 1* and CHEN Hui-Peng 1* 1.Institute of Biotechnology,Academy of Military Medical Sciences, Beijing 10 0071, China 2.College of Life Science, Jilin University, Changchun 130023,China  
The lysin gene of Bacillus anthracis-diagnosing bacteriophage,obtaine d by PCR amplification,was cloned into the Escherichia coli exepression vect or p ET22b which has been cleaved by EcoRⅠ and NdeⅠ.The recombinant vector pET22b-γ l ysin was verified to be correctly constructed by PCR, sequencing and enzyme dige stion,and highly expressed in E.coli BL21(DE3), which accounted for about 40 petcent of total protein in E.coli BL21(DE3),while in the 5L fermentor the ex pres sion level reached 15g/L.After expression, disruption and purification with thre e-step chromatography, Streamline SP, SP HP and Sephacryl S-100,the recombinan t γ lysin was finally obtained with purity of higher than 95 percent as determ ine d by gel scan. The final yield following SP HP was 19.1 percent,with a greater - than-350-fold increase in specific activity.The pure enzyme has been shown act iv e to Bacillus anthracis,and not to E.coli,Bacillus subtilis and Bacill us cereus. Its specific activity was about 1400 u/mg.
【CateGory Index】: R346
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Chinese Journal Full-text Database 3 Hits
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