Design and Construction of Human Tissue Plasminogen Activator Expression Vector for The White Button Mushroom, Agaricus bisporus

The similarity of the white button mushroom glycosylation to mammalian cells, caused a significant position to expression of recombinant glycosylated proteins in this mushroom. Human t-PA is a glycosylated protein that degrades the fibrin network formed within blood clots. The gpdII promoter of the button white mushroom strains IM008 and Holland737 were isolated via PCR and the gpdII promoter of the strain IM008 inserted before of hph gene in pCAMBIA1304 plasmid and pCAMBIAH8 constructed. The gLI7 and gLI8 fragments (resulted from connectivity of gpdII promoter of strains Holland737 and IM008, respectively to intron number seven of gpdII gene and laccase1 gene signal peptide) inserted to pCAMBIAH8 and then p13H88 and p13H87 created. Finally the tPA gene cloned to pCAMBIAH88 and pCAMBIAH87 and then p13H88T and p13H87T constructed, respectively. Constructs in this research can be used for tPA production as well as for genetic improvement program of the white button mushroom.