摘要
目的构建长链非编码RNA-TP53TG1的真核表达克隆,并探索其在神经胶质瘤中对糖剥夺应激反应的影响。方法用RT-PCR方法从人胶质瘤细胞中扩增出TP53TG1;构建了TP53TG1的全长真核表达克隆;实时定量PCR检测其在U87MG细胞内的表达;同时用低糖(0.3 g/L葡萄糖、8 h)处理;real-time PCR检测GRP78、IDH1和PKM2的表达水平。结果成功构建了真核重组表达质粒pCIG-TP53TG1;在转染U87MG细胞36 h后,可见绿色荧光的表达,U87MG细胞中TP53TG1 mRNA升高了2.9×106倍(P<0.05);过表达TP53TG1的同时低糖处理,GRP78和IDH1 mRNA的表达水平显著升高(P<0.05),而PKM2 mRNA的表达水平显著降低(P<0.05)。结论在U87MG细胞中,TP53TG1可能通过影响GRP78、IDH1和PKM2 mRNA的表达,而参与到对糖剥夺的应激反应过程。
Objective To construct full length clone of TP53TG1 into eukaryotic expression vector, and to explore its overexpression effect on glucose deprivation stress response. Methods TP53TG1 was amplified from human gli oma cell by RT-PCR, and the eukaryotic expression clone of TP53TG1 was constructed. Then, its expression in U87MG cells was detected by real time PCR. Furthermore, we overexpressed TP53TG1 and meanwhile treated with low glucose （0. 3 g/L,8 h） in U87MG cells, and measured the expression of GRP78, IDH1 and PKM2 mRNA by real time PCR. Results TP53TG1 eukaryotic expression clone was successfully constructed. After the clone was transfected into U87MG cells for 36 hours, green fluorescence was seen. The expression of TP53TG1 was increased by 2. 9 x 106 times in U87MG cells （P 〈0. 05）. As a result of over expression of TP53TG1 and low glucose treat ment simultaneously in U87MG cells, GRP78 and IDHI mRNA expression were significantly increased （P 〈 0. 05 ）, while PKM2 mRNA significantly reduced （ P 〈 0. 05 ）. Conclusions TP53TG1 may be involved in the stress response of U87MG cells under glucose deprivation through influencing the expression of GRP78, IDHI and PKM2 mRNA.
出处
《基础医学与临床》
CSCD
北大核心
2013年第6期680-684,共5页
Basic and Clinical Medicine
基金
国家自然科学基金(31071203)
