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Cloning and Expression of the cDNA of a Murine Soluble Fas

Cloning and Expression of the cDNA of a Murine Soluble Fas
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摘要 In order to regulate the apoptosis induced by Fas FasL system, a soluble isoform of mouse Fas was cloned from thymocytes of immature mice with the primers designed according to the full length Fas cDNA sequence in the GeneBank. It was directionally inserted into the intermedium vector pUC19. DNA sequencing proved that it was consistent with the expected sequence. Then it was subcloned into the eukaryotic expression vector pCA13, which was used to construct the recombinant vector pCA13 FasC. By lipofectamine (LF2000) mediated transfection, pCA13 FasC was transfected into the 293 cells. RT PCR and Western blot indicated that the murine soluble Fas C protein was expressed in the 293 cells. Apoptosis inducing test showed that the expression of this murine Fas C could block the Fas induced apoptosis, which confirmed the biological activity of the recombinant Fas C. In order to regulate the apoptosis induced by Fas FasL system, a soluble isoform of mouse Fas was cloned from thymocytes of immature mice with the primers designed according to the full length Fas cDNA sequence in the GeneBank. It was directionally inserted into the intermedium vector pUC19. DNA sequencing proved that it was consistent with the expected sequence. Then it was subcloned into the eukaryotic expression vector pCA13, which was used to construct the recombinant vector pCA13 FasC. By lipofectamine (LF2000) mediated transfection, pCA13 FasC was transfected into the 293 cells. RT PCR and Western blot indicated that the murine soluble Fas C protein was expressed in the 293 cells. Apoptosis inducing test showed that the expression of this murine Fas C could block the Fas induced apoptosis, which confirmed the biological activity of the recombinant Fas C.
出处 《Journal of Huazhong University of Science and Technology(Medical Sciences)》 SCIE CAS 2002年第3期186-189,196,共5页 华中科技大学学报(医学英德文版)
基金 a grant from the NationalNatural Science Fundation of China(Serial No.3 9970 70 8)
关键词 soluble Fas MOUSE eukaryotic expression soluble Fas mouse eukaryotic expression
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参考文献5

  • 1Cascino I,Fiucci G,Papoff G et al.Three functional soluble forms of the human apoptosis-inducing Fas molecule are produced by alternative splicing[].J Immunol.1995
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  • 3Walker P R,Saas P,Dietrich P.Role of Fas ligand( CD95L ) in immune escape: the tumor cell strikes back[].J Immunol.1997
  • 4Watanabe-Fukunaga R,Brannan C I,Itoh N et al.The cDNA structure, expression, and chromosomal assignment of the mouse Fas antigen[].J Immunol.1992
  • 5Hughes D P,Crispe I N.A naturally occurring soluble isoform of murine Fas generated by alternative splicing[].The Journal of Experimental Medicine.1995

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