siRNA靶向EDAG-1基因对K562细胞株生长的影响

背景与目的：胚胎发育相关基因-1（EDAG-1）是造血系统发育相关的新基因，定位于9q22。本研究探讨抑制EDAG-1基因的表达对白血病细胞株生长的影响及其机制。方法：将靶向EDAG-1基因的特异序列连接到逆转录病毒载体中，脂质体介导的重组质粒转染包装病毒细胞（PT67），收集细胞上清感染K562细胞株，经嘌呤霉素筛选稳定抑制EDAG-1基因表达的细胞株，RT-PCR检测EDAG-1基因的抑制情况，应用MTT、流式细胞仪检测转染细胞的增殖能力及细胞周期。结果：成功构建了靶向EDAG-1基因的逆转录病毒重组质粒，建立了抑制EDAG-1基因表达的K562细胞株。抑制EDAG-1基因表达的细胞株增殖速度、增殖指数（PI）、增殖活性（SPF）相应降低，其G0／G1期细胞比例明显升高。结论：EDAG-1基因的沉默可以抑制白血病细胞株的增殖，使其阻滞在G0／G1期，提示EDAG-1基因可能是白血病治疗的一个有效靶点。

Impact of siRNA targeting for EDAG-1 on growth of K562 cell line

Background and purpose:Embryonic developmentassociated gene-1(EDAG-1) locates in chromosome 9q22,which is considered to be a human hematopoiesis-specific gene.The purpose of this study was to explore the effects of silent EDAG-1 by siRNA on the growth of leukemia cell line and its molecular mechanism.Methods:The oligonucleotides for EDAG-1 were cloned into retroviral vector,subsequently the recombinant retroviral vectors based RNAi were transfected into packing cell line.K562 was infected by the virus supernatant collected from packing cell line,the stable cell lines were selected with puromycin.The reduction of EDAG-1 was inspected with RT-PCR;and proliferation and cell cycle were assayed with MTT and flow cytometry respectively.Results:The recombinant retroviral vectors were successfully constructed;the stable K562 cell lines with a persistent knockdown of EDAG-1 were established.The proliferation of cell lines was significantly inhibited after silence of EDAG-1.SPF and PI were reduced,G_(0)/G_(1) arrest was observed in the cell line with the silence of EDAG-1.Conclusions:Inhibition of the expression of EDAG-1 can reduce the proliferation of leukemia cell lines,and arrested them in G_(0)/G_(1) phase.The primary results suggest that EDAG-1 might serve as an effective target for the treatment of leukemia.