Cloning,High Level Expression and Purification of Porcine IFNγ

Peripheral blood lymphocytes from a single swine were stimulated with Concavadin A for 17h,and the total RNA was isolated from it.Then,the mRNA specific for porcine IFNγ was amplified by reverse transcription polymerase chain reaction.After sequencing,the IFNγ gene has been successfully inserted into vector pJLA\|503 and highly expressed in E.coli. Recombinant porcine IFNγ expressed as inclusion body,which was dissolved in 7mol/L guanidine chloride and subsequently renatured by dilution in refolding buffer containing 0.5mol/L L\|arginine.In order to obtain pure protein,the renatured IFNγ was pruified by the chromatographies of SP\|Sepharose FF and Sephacryl S\|200 HR.As a result,the final pure product can been seen as a single band in SDS\|PAGE, and the cytokine activity was verified by inhibiting the cytopathic effect.