hpve7通过下调microrna-17抵抗化疗药物作用的机制分析word论文.docxVIP

AbstractObjective: The purpose of this study is to discurss chemoresistance mechanisms caused by human papillomavirus (HPV) from the perspective of miRNA.Methods:(1)MTT assay detected the inhibition rates of docetaxel in different concentrations on two stably transfected PC3; (2)The expression level of the genes relating to cell cycle; (3) The expression level of candidate miRNA was detected by RT-PCR; (4) Analysis software predicted the potential target genes of the candidate miRNA; (5) Western blot detected the protein expression level of potential target gene; (6) The construction of recombinant luciferase reporter gene plasmid pGL3-CM / miRNA-17-CDK6 and it’s function analysis; (7) Function analysis of miRNA analogues / inhibitor (Mimics / Inhibitors) .Results:(1) MTT assay suggested that the inhibition rate of docetaxel for PC3 cells with pcDNA3.1(-)-E7 is lower than PC3 cell with pcDNA3.1(-); (2) miRNA-17 was elected to be the candidate miRNA, and it was reduced significantly after E7 transfection, increased significantly after dosing in PC3 cell transfected with pcDNA3.1(-), while there was no statistical difference after dosing in PC3 cells with pcDNA3.1(-)-E7; (3) CDK6 has been predicted to be a potential target gene by computer analysis software; (4) The CDK6 mRNA was upregulated significantly after E7 transfection, downregulated significantly after dosing in PC3 cell with pcDNA3.1(-), however, there was no statistical difference after dosing in PC3 cells with pcDNA3.1(-)-E7; (5) The protein expression levels of CDK6 detected by western blot were consistent to the expression levels of CDK6 mRNA; (6) The construction of recombinant luciferase reporter gene plasmid was successful. recombinant plasmid was transfected into the two kinds of stably transfected cells, and treated with drug, then detected their luciferase activity. Luciferase activity increased significantly after E7 transfection, reduced significantly after dosing in PC3 with pcDNA3.1(-), there