Construction of the eukaryotic expressing vector of PcDNA3.1 rhGM-CSF and its effect on the proliferation and differentiation of K562 cells / 中国免疫学杂志

ABSTRACT

Objective:To construct eukaryotic expressing vector of PcDNA3.1-rhGM-CSF and to investigate its effects of suppressing growth and inducing differentiation in K562 cells.Methods:The eukaryotic expressing vector PcDNA3.1-hGM-CSF was constructed by means of PCR and T-A clone techniques as well as directional cloning techniques.It was affirmed by the restriction map and DNA sequence analysis,and then transfected into K562 cells.It was observed that the target gene of the recombinant vector was expressed and exerted in K562 cells 72 h later,RT-PCR was used to affirm the expression of rhGM-CSF in K562 cells;Cell morphological,cell proliferation assay and immunohistochemistry were used to observe the effect of PcDNA3.1-rhGM-CSF on the growth and differentiation of K562 cells.Results:①The recombinant eukaryotic expressing vector PcDNA3.1-GM-CSF was constructed successfully;②Recombinant GM-CSF was expressed in the transfected K562 cells and the transfected K562 cells could differentiate into monocyte and macrophage cells;③ The proliferation of K562 cells was suppressed.Conclusion:The recombinant eukaryotic expression vector PcDNA3.1-rhGM-CSF was constructed successfully;Transfected K562 cell could differentiate into monocyte and macrophage cell.