CAJ | 학술논문

目的构建微小RNA let-7c重组慢病毒载体,验证转染该载体系统后对肝癌细胞增殖的影响方法构建表达含494bp let-7c基因的重组慢病毒载体,检测let-7c及对照组病毒滴度分别为5.01×10E8 TU/m1、5.15×10E8TU/ml;转染HepG2细胞,以荧光定量聚合酶链反应(PCR)对let-7e表达水平进行检测；细胞计数试剂盒(CCK-8)试验评价let-7c过表达后对HepG2细胞增殖的影响.结果构建的let-7c慢病毒载体经质粒酶切和测序鉴定正确；转染HepG2细胞72 h后,let-7c表达上调312倍；CCK-8法检测显示HepG2细胞增殖受到明显抑制(P＜0.01).结论成功构建Iet-7c重组慢病毒载体并感染HepG2细胞后高效表达成熟let-7c,抑制了HepG2肝癌细胞的增殖.
목적 구건미소RNA let-7c중조만병독재체,험증전염해재체계통후대간암세포증식적영향 방법 구건표체함494bp let-7c기인적중조만병독재체,검측let-7c급대조조병독적도분별위5.01×10E8 TU/m1、5.15×10E8TU/ml;전염HepG2세포,이형광정량취합매련반응(PCR)대let-7e표체수평진행검측；세포계수시제합(CCK-8)시험평개let-7c과표체후대HepG2세포증식적영향.결과 구건적let-7c만병독재체경질립매절화측서감정정학；전염HepG2세포72 h후,let-7c표체상조312배；CCK-8법검측현시HepG2세포증식수도명현억제(P＜0.01).결론 성공구건Iet-7c중조만병독재체병감염HepG2세포후고효표체성숙let-7c,억제료HepG2간암세포적증식.
Objective To construct recombinant lentiviral vector of miRNA let-7c and verify its effect on liver cancer cell proliferation.Methods Recombinant lentiviral vector containing 494 bp let-7c gene was constructed.The titers of Let-7c and the controls were 5.01 × 10E8 and 5.15 × 10E8 TU/ml respectively.Let-7c lentiviral vector was transfected into HepG2,and the expression of let-7c was detected by using real time polymerase chain reaction (PCR).Cell counting Kit-8 (CCK-8) method was used to testify HepG2 cell proliferation after the vector transfection.Results The successful construction of let-7c lentiviral vector was confirmed by plasmid enzyme digestion and DNA sequencing.Seventy-two h after the vector transfection,the expression of mature let-7c was increased by about 312 folds,compared to blank vector transfection.The over-expression of let-7e significantly suppressed HepG2 cell prolifieration (P ＜0.01 ).Conclusion The let-7c recombinant lentiviral vector was successfully constructed.After the vector was transfected to HepG2 cells,mature let-7c was effectively expressed and suppressed the HepG2 liver cancer cell proliferation.