Transient expression of human thrombopoietin in cell directed by rat WAP promotor

ObjectiveTo study the role of 5'untranslated region(UTR) and introns in human thrombopoietin (TPO) transient expression through TPO eukaryotic expression vectors directed by rat whey acidic protein(WAP) promoter.MethodsSix TPO eukaryotic expression vectors directed by rat WAP were constructed with gene engineering technology and identified by restriction enzyme and sequencing.The recombinant plasmids were transfected into HC-11 and Cos-1 cells.Forty-eight hours after the transfection, the expression of TPO in supernatant was analyzed with sandwich enzyme-linked immunosorbent assay (ELISA).ResultsThe recombinant plasmids were constructed as designed.The expression level of 6 recombinant plasmids in HC-11 cells was higher than that in COS-1 cells with the order of pTPOWE,pTPOWA,pTPOWC,pTPOWB,pTPOWD,and pTPOWF from high to low expression.The recombinant plasmid containing intronⅤ showed the highest expression level and the plasmid harboring TPO genome expressed in the lowest level.ConclusionThe intron Ⅴ may contain specific sequence enhancing TPO expression and the TPO genome may contain inhibitory elements for TPO expression.