Construction of human single chain interleukin-12 fusion gene by recombinant PCR
Abstract：Objective To study biological functions of single chain IL-12protein，we constructed fusion gene of recombinant human single chain IL-12（hscIL-12） and expressed the fusion gene in mammalian cell．Methods A hydrophobic polypeptide linker （Gly4Ser）3 was used to splice two different gene fragments of p40 and p35 subunits of hIL-12 （hIL-12 p40-linker-p35） by recombinant PCR for constructing rhscIL-12 fusion gene in vitro．After sequencing，rhscIL-12 fusion gene was inserted into pcDNA3．1（＋） eukaryotic expressing plasmid and then transfected into COS-7 cell by lipofection．The fusion protein expressed was analysed with Western blot．Results Sequence analysis showed that the splicing order，the direction and the sequence in hscIL-12 fusion gene were completely correct．Expressed hscIL-12 fusion protein in COS-7 cells combined with mouse anti-human IL-12 monoclonal antibody specifically．Western blot showed the appearance of 70 000 u band of fusion protein．Conclusion The result indicates that recombinant PCR is a very effective and reliable method for gene recombination in vitro．
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Publication Date：2001-01-01
Online Publishing Date：2025-08-15(First online date of this platform, not the publication date of the document)
Pages：5( 22-26 )