Construction and expression of eukaryotic expression vectors for mouse macrophage Ipr1 gene

To construct mouse Ipr1 gene (intracellular pathogen resistance 1)eukaryotic expression vector,the DNA fragment of Ipr1 gene was obtained through RT-PCR method from C57BL/6J mouse thymus.The gene fragment were cloned into simple vectors pMD19-T and transformed into E.Coli JM109.and the recombinant plasmid was identified by PCR,restrict endonuclease digestion and sequencing.Ipr1 gene was subcloned into pQE-Trisystem and the recombinant plasmid pQE-Trisystem-Ipr1 was comfirmed by restrict endonuclease digestion.Then the pQE-Trisystem-Ipr1 was transfected into RAW264.7,after transient transfection expression product of Ipr1 gene was detected by RT-PCR.Thus,the whole coding sequence of Ipr1 gene was successfully cloned as expected.A point mutation was found by sequencing.After the point mutation was repaired and the accurate recombinant pQE-Trisystem-Ipr1 was obtained,Ipr1 gene expressed in transfected RAW264.7 was detected by RT-PCR.Through these procedures,the coding sequence of Ipr1 gene was obtained from C57BL/6J mouse thymus and the eukaryotic expression vector for Ipr1 gene was successfully constructed.This laid the foundation for the further stndy on the funetions of Ipr1 gene.