含EWS-FLI1结合序列的DTA载体对报告基因表达抑制的研究

DTA vector containing the binding sequence of EWS-FLI1 depresses the activation of luciferase

目的 研究含尤文家族肿瘤EWS FLI1结合序列的DTA载体对报告基因表达的抑制作用。方法 构建含有EWS FLI1结合序列和白喉毒素A链基因的表达载体 pS2 DTA。共同转染不同剂量梯度的 pS2 DTA和pS2到尤文肉瘤细胞和对照细胞 ,检测荧光强度。结果 每增加一个 pS2 DTA剂量梯度 ,在尤文肉瘤细胞中都可以检测到虫荧光素酶的表达显著降低 ,而在对照细胞中这种作用却不明显 ;在同一个pS2 DTA剂量梯度下 ,虫荧光素酶在尤文肉瘤细胞的表达都显著高于非尤文肉瘤细胞。结论 pS2 DTA转染可以抑制尤文肉瘤细胞内其它基因表达。

Objective To study the DTA vector containing the Ewing family of tumor's EWS-FLI1 binding sequence depresses the co-transferred luciferase reporter plasmid.Methods To construct the DTA expression vector (pS2-DTA), the luciferase gene in pS2 was replaced with DTA gene at the points of NcoⅠand XbaⅠ by molecular technique. To compare the activation of luciferase, pS2 and pS2-DTA were co-transferred into cultured Ewing's sarcoma cell (SK-ES) and control cell (Hela).Results The luciferase activity in Ewing's sarcoma cells was significantly depressed, with the high concentration of pS2-DTA. The expression of luciferase is higher in SK-ES than that in Hela at the same concentration.Conclusions The pS2-DTA can depress the activation of luciferase, and this function is stronger when concentration of pS2-DTA co-transferred.