引用本文:姚 莉 1,孙 艳1,查 何1,彭惠民2.miRNA-451表达质粒的构建及对人A549肺癌细胞侵袭转移及凋亡的影响[J].重庆医科大学学报,2012,37(10):841~845
miRNA-451表达质粒的构建及对人A549肺癌细胞侵袭转移及凋亡的影响
Construction of miRNA-451 expression plasmid and its effect on invasion,metastasis and apoptosis in human A549 lung cancer cells
DOI:
中文关键词:  肺癌  miRNA- 451  侵袭  转移  凋亡
英文关键词:lung cancer  miRNA-451  invasion  metastasis  apoptosis
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作者单位
姚 莉 1,孙 艳1,查 何1,彭惠民2 (重庆医科大学 1. 细胞生物学及遗传学教研室2. 分子医学与肿瘤研究中心重庆 400016) 
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中文摘要:
      目的:探讨miRNA-451对人A549肺癌细胞侵袭转移及凋亡的影响。方法:Hsa-miRNA-451序列通过miRBase数据库查找,根据此序列设计出2条寡核苷酸片段,将目的基因经过退火处理后克隆至真核表达载体pGenesil-1.1质粒中,从而构建成为miRNA-451真核表达载体(pGenesil-1.1-miRNA-451 质粒)。将 pGenesil-1.1-miRNA- 451 及pGenesil-1.1-control 质粒分别转染人A549肺癌细胞,经过G418的筛选,建立pGenesil-1.1-miRNA-451及pGenesil-1.1-control 稳定表达的细胞株。流式细胞术分析检测细胞凋亡的情况;Transwell小室检测miRNA-451对细胞侵袭能力的影响;细胞划痕实验检测细胞的迁移情况。结果:经测序证实miRNA-451真核表达质粒构建成功,与未处理的人A549肺癌细胞和稳定表达 pGenesil-1.1-control 的细胞相比,稳定表达了pGenesil-1-miRNA-451的人A549肺癌细胞的侵袭和迁移能力均明显受到抑制影响,而细胞凋亡无明显变化(P >0.05)。结论:miRNA-451对人A549肺癌细胞侵袭转移具有显著的抑制作用,但对细胞凋亡作用不明显。
英文摘要:
      Objective:To explore of the effect of miRNA-451 on invasion,metastasis and apoptosis of human A549 lung cancer cells. Methods:Hsa-miRNA-451 sequence was found in miRBase database and two oligonucleotides were designed according to the se-quence. The targeted gene was annealed and cloned to eukaryotic expression vector plasmid pGenesil-1.1 thus miRNA-451 eukaryotic expression vector was built(pGenesil-1.1-of miRNA-451 plasmid). PGenesil-1.1-miRNA-451 and pGenesil-1.1-control plasmid were transfected into human A549 lung cancer cells and stably expressed pGenesil-1.1-miRNA-451 and pGenesil-1.1-control cell lines were established after being selected by G418. Flow cytometry was employed to detect cell apoptosis,Transwell chamber to detect miRNA-451 cell invasion and cell scratch assay to detect cell metastasis. Results:Sequencing showed that miRNA-451 eukary-otic expression plasmids were successfully constructed. Cell invasion and metastasis were inhibited in stably expressed pGenesil-1.1-miRNA-451 cell lines compared with those in untreated human A549 lung cancer cells and stably expressed pGenesil-1.1-control cell lines,however,there was no change in cell apoptosis(P >0.05). Conclusion:miRNA-451 has significant inhibitory effect on in-vasion and metastasis of human A549 lung cancer cell,however,not on cell apoptosis.
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