Title: Cloning and Expression of EndoglucanaseI of T.Knoningii
Abstract: Objective:To construct Prokaryotic expression vector of endoglucanaseⅠ and study the activity of the expressed production.Method:The gene coding endoglucanaseⅠ was amplified from total RNA of T.Knoningii by RT-PCR technique.The PCR product was cloned into the expression Vector pET·His which is controlled by T7promoter,and the prokaryotic expression vector pET·His-EGⅠ was thus constructed successfully.The reconstructed plasmid was transformed into E.coli BL21(DE3) plysS competent cel1.The bacterium was induced by IPTG(0.4 mmol/L)and analyzed by 13%SDS-PAGE.Result:Approximately 45kDa exogenous protein was observed on the SDS-PAGE.The result showed the enzyme optimal activity at pH 6.0 and 30℃-40℃,and the recombinant enzyme was obviously activated by Mn2+ ion.Conclusion:EGⅠ protein was expressed in E.coli and showed the activity,which laid a foundation of EGⅠfurther biology research.
Publication Year: 2008
Publication Date: 2008-01-01
Language: en
Type: article
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