RNA干扰MeCP2对肺癌细胞A549增殖的影响

Effects of Inhibition of Methyl-CpG-inding Protein 2 by RNA Interference on Proliferation of Lung Carcinoma Cell A549

目的研究甲基化结合蛋白MeCP2对肺癌细胞A549增殖的影响。方法针对GeneBank中MeCP2基因的不同靶点,设计并合成两条DNA干扰序列,克隆到真核表达载体pSilencer-2.1-U6,构建能够转录形成MeCP2-shRNA的重组质粒pSilencer-MeCP2,经PCR、测序鉴定正确后,稳定转染A549细胞,在mRNA和蛋白水平观察两重组质粒对MeCP2的干扰作用,MTT法检测对细胞增殖能力的影响。结果成功构建两个针对MeCP2不同靶序列的shRNA真核表达质粒,两质粒在mRNA和蛋白水平可以使MeCP2表达明显下调,稳定转染两质粒的A549细胞增殖能力明显减弱。结论甲基化结合蛋白MeCP2参与了肺癌细胞的恶性增殖。

Objective To study the effects of MeCP2 in proliferation of lung cancer cell lines A-549. Methods By the sequence of human MeCP2 gene in OeneBank, the oligonucleotides encoding shRNA targe- ting the MeCP2 gene were designed and sythesized, then directionally cloned into plasimd pSliencer-2, 1- U6, confirmed by PCR and DNA sequencing analysis, the recombinant plasimd was transfected into A549 cell line by lipofectamine TM2000. The expression of MeCP2 at mRNA and protein was examined by RT-PCR and Western blot respectively, the changes of cell proliferation were observed by MTT assay. Results The two shRNAs expressing plasmids targeting the MeCP2 gene were constructed and transfected into A549 cell line successfully. The expression of MeCP2 in mRNA and protein levels was reduced visibly. The down regulation of MeCP2 inhibited the proliferation of A549 cell significantly. Conclusion The MeCP2 protein plays an important role in the proliferation of lung cancer cell.