title: Construction and identification of a human liver specific microRNA eukaryotic expression vector.
abstract: MiR-122 is one of the non-coding RNAs which showed its effects on the lipo-metablism, virus infection and HCC forming through regulation of liver gene expression. Its eukaryotic expression vector was constructed by using pSuper which was widely applied in the siRNA expression. The precursor of human miR-122 gene was amplified by polymerase chain reaction (PCR) from the human genomic DNA. The positive clones were screened by PCR and restriction enzyme digestion. The new expression vector of miR-122 was named pHsa-m122. PHsa-m122 and its controls were transfected to HepG2 cells. The miR-122 expression activity was evaluated by GFP122i sensor reporter plasmid through fluorescence detection and Western blot. It was shown that the fluorescence intensity of GFP122si and pHsa-m122 co-transfection group was weaker than that of the controls, so the functional activity of expressed miR-122 was detected. When HepG2 cells were co-transfected with HBV1.3 and pHsa-m122 plasmids, the results showed miR-122 may down-regulate the gene expression of HBV. The human liver specific microRNA eukaryotic expression vector of miR-122 was constructed successfully, which may facilitate further study of its function in the development of liver virus infection diseases and HCC.
authors.affiliation: Department of Pathogenic Biology, Tongji Medical College of Huazhong University of Science & Technology, and Department of Infectious Diseases, Tongji Hospital, Wuhan 430030, China.
authors.forename: Shanshan
authors.initials: S
authors.lastname: Chen
authors.forename: Ming
authors.initials: M
authors.lastname: Ni
authors.forename: Bing
authors.initials: B
authors.lastname: Yu
authors.forename: Tingting
authors.initials: T
authors.lastname: Lv
authors.forename: Mengji
authors.initials: M
authors.lastname: Lu
authors.forename: Feili
authors.initials: F
authors.lastname: Gong
mesh_terms: D000951:Antigens, Neoplasm / Q000235:genetics; D006528:Carcinoma, Hepatocellular / Q000235:genetics / Q000628:therapy; D002460:Cell Line; D045744:Cell Line, Tumor; D004797:Enzyme-Linked Immunosorbent Assay; D015967:Gene Expression Regulation, Viral; D015316:Genetic Therapy; D005822:Genetic Vectors / Q000737:chemistry; D049452:Green Fluorescent Proteins; D006509:Hepatitis B / Q000235:genetics / Q000628:therapy; D006511:Hepatitis B Antigens / Q000235:genetics*; D006515:Hepatitis B virus / Q000235:genetics / Q000276:immunology; D006801:Humans; D050356:Lipid Metabolism / Q000235:genetics; D008099:Liver / Q000378:metabolism / Q000821:virology; D035683:MicroRNAs / Q000235:genetics* / Q000378:metabolism / Q000627:therapeutic use; D009928:Organ Specificity; D014162:Transfection
chemical_list: D000951:Antigens, Neoplasm; D006511:Hepatitis B Antigens; D035683:MicroRNAs; D049452:Green Fluorescent Proteins