Title: Expression of hCD74 truncated fragment in insect baculovirus vector system
Abstract: Objective To establish an expression system of human Ⅱ CD74 extracellular fragment by insect baculovirus vector expression system in insect cells.Methods hCD74 full extracellular fragment(73~232 aa) and the core area combined with MIF(109~192 aa) were amplified by PCR,and inserted into pFastBacDual vector placed promoter of PPH.Recombinant plasmid was transformed into E.coli DH10Bac and obtained recombinant virus bacmid by site specific transposition.Recombinant bacmid was transfected into Sf-21 cell line.The target mRNA transcription was tested by RT-PCR and recombinant protein expression was analyzed by SDS-PAGE and Western blot.Results The full extracellular fragment(73~232 aa) of hCD74 expressed in insect cells and the target protein was detected using antibody against His tag and CD74.In addition,it achieved the maximum expression in cells after 72 h infection and inoculation using the virus titer MOI 4.The core area of fragment(109~192 aa) was positive for mRNA transcription,but not protein expression.Conclusion hCD74 extracellular fragment is successfully expressed in Sf-21 cell,and the protein expression conditions are optimized.
Publication Year: 2010
Publication Date: 2010-01-01
Language: en
Type: article
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