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High Expression and Purification of IFN-β1b and Its Antiviral Activity

LI Wu ping,LV Hong liang,DUAN Zhao jun,ZHANG Li ping,LIU Yong qing,WU Bin wen, CHEN Yong,ZHANG Cheng hai,YI Zhuo an,WEI Kai kun,HOU Yun de (National Institute for Viral Disease Control and Prevention,China CDC,Beijing 10005  
IFN-β1b,an analogue of human IFN-β, where serine was genetically engineered to substitute for cysteine at position 17,was produced in E.coli.In order to obtain high expression,we synthesized the gene of IFN-β-1b artificially using the preferred codons of E.coli,and inserted it into plasmid pBV220,and transformed to E.coli DH5α.The processes of IFN-β1b manufacturing consisted of fermentation and a series of purification steps.Briefly,a production strain of E.coli harboring the modified IFN-β gene was grown in fermenter.The synthesis of IFN-β was under the control of inducible promoter P RP L.The newly synthesized IFN-β was deposited in bacterial cells as inclusion bodies.The culture was harvested and bacterial cells were disruptd to release the intracellular IFN-β1b.IFN-β1b was then solubilized using a buffer solution containing sodium dodecyl sulphate(SDS)and reduced using dithiothreitol(DTT).Purification processes consisted of organic extraction,size exclusion chromatography,desalting,oxidation to form correct disulphide bond followed by reverse phase chromatography and rotary vaporization.The antiviral activity of IFN-β-1b on cell lines of different species was different.
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