Construction of miR-31 lentiviral vector and its effect on proliferation and migration of cervical cancer HeLa cells

WANG Linlin; HU Xiaoxia; LIU Fei

doi:10.3969/j.issn.1002-266X.2017.28.001

Abstract：Objective To construct the miR-31 lentiviral vector and to investigate its effect on proliferation and migration of cervical cancer HeLa cells.Methods PCR was used to amplify the target gene which included gene miR-31.The target gene was designed with appropriate enzyme sites at the end of the primer pairs.We inserted the PCR products into linearization of lentiviral vector after the double digestion.Restriction endonuclease analysis and DNA sequencing had confirmed the sequence of the recombinant plasmid pCDH-CMV-MCS-EF1-copGFP-miR-31.HeLa cells were co-transfected with the lentiviral vectors pCDH-CMV-MCS-EF1-copGFP-miR-31, pPACKH1-GAG, pPACKH1-REV and pVSV-G, the titer of the lentivirus was detected.HeLa cells in the logarithmic phase were randomly divided into the control group without any treatment, the miR-31 group treated with lentivirus miR-31, and the negative control group given empty lentivirus.The qPCR was used to detect the expression of miR-31 in HeLa cells of each group.The cell proliferation was detected by clone formation assay and MTT assay, and the cell migration ability was detected by Transwell assay.Results The recombinant lentivirus expression plasmid was constructed successfully, with lentivirus titer of 3×107 TU/mL.The expression of miR-31 in HeLa cells of the miR-31 group was significantly higher than that of the other two groups (all P<0.05);no significant difference was found between the control group and the negative control group (P>0.05).Compared with the other two groups, the proliferation and migration abilities of HeLa cells were increased in the miR-31 group (all P<0.05);no significant difference was found between the control group and the negative control group (P>0.05).Conclusion The miR-31 lentivirus expression plasmid is constructed successfully, and the proliferation and migration abilities of HeLa cells with stale expression of miR-31 are significantly enhanced.