人LncRNA MIR31HG基因稳定转染PC9细胞系的构建及其对细胞增殖和迁移的影响

doi:10.13481/j.1671-587x.20190410

Abstract

Abstract: Objective:To detect the effect of over-expressed human MIR31HG gene on the proliferation and migration of PC9 cells, and to clarify the mechanism of oncogene MIR31HG. Methods:The full-length sequence of long non-coding RNA (LncRNA) MIR31HG was amplified by RT-PCR and cloned into the pcDNA3.1(-) eukaryotic expression vector. The PC9 cells were transfected with the pcDNA3.1-MIR31HG overexpression vector and control vector pcDNA3.1. The construction of MIR31HG overexpression vector was detected by enzymatic digestion identification. The stable cell lines with overexpression of MIR31HG (PC9-pcDNA3.1-MIR31HG,stable transfection group) and control cell lines (PC9-pcDNA3.1,empty vector group) were established by G418 drug screening, and the expression level of MIR31HG gene in stably transfected cell line was detected by RT-PCR. CCK-8 method and scratch healing assay were used to detect the proliferation activities and migration abilities of PC9 cells. Results:The agarose gel electrophoresis results showed that the specific gene fragment of MIR31HG was obtained by amplification successfully. The gene fragments of target gene and vector were produced by double enzyme digestion of MIR31HG eukaryotic expression vector. The RT-PCR results showed that the MIR31HG RNA expression level in the cells in stable transfection group was significantly higher than that in empty vector group(P<0.05).The results of CCK-8 test showed that the proliferation activities of the cells in stable transfection group were significantly higher than those in empty vector group at 24, 36 and 48 h after culture(P<0.01).The results of scratch healing assay showed that the scratch healing rate of cells in stable transfection group was significantly higher than that in empty vector group at 48 h after culture(P<0.05). Conclusion:The eukaryotic overexpression vector and the PC9 cell line stably transfected with human LncRNA MIR31HG gene are constructed successfully, and MIR31HG overexpression can promote the proliferation and migration of lung cancer PC9 cells.

Key words: long non-coding RNA, MIR31HG, PC9 cell lines, cell proliferation, cell migration

CLC Number:

DAI Juanjuan, YANG Lijuan, DU Jing, MIAO Shuang, XI Sichuan, LI Chen, WU Yan. Construction of PC9 cell lines stably transfectedwith human LncRNAMIR31HG gene and its effects on cell proliferation and migration[J].Journal of Jilin University(Medicine Edition), 2019, 45(04): 801-806.

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Construction of PC9 cell lines stably transfectedwith human LncRNAMIR31HG gene and its effects on cell proliferation and migration

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