title: Methods for analyzing microRNA expression and function during osteogenic differentiation of human adipose tissue-derived mesenchymal stem cells.
abstract: MicroRNAs (miRNA) are single-stranded RNA molecules of 21-23 nucleotides in length that regulate gene expression at the posttranscriptional level. They may play important roles during osteogenic differentiation of adipose tissue-derived mesenchymal stem cells (hASC). In this chapter, we focus on the methods and strategies for elucidating miRNA function during osteogenic differentiation. We describe a miRNA expression analysis protocol, and a lentiviral vector strategy for the ectopic expression of miRNA in hASC to determine the role of miRNA during osteogenic differentiation. We also describe miRNA inhibition to further determine the role of miRNA during osteogenic differentiation, and a luciferase assay to demonstrate direct binding between a specific miRNA and its putative target.
authors.affiliation: Department of Physiology, School of Medicine, Pusan National University, Gyeongsangnam-do, Beomeo-ri, Korea.
authors.forename: Yeon Jeong
authors.initials: YJ
authors.lastname: Kim
authors.forename: Jin Sup
authors.initials: JS
authors.lastname: Jung
mesh_terms: D000273:Adipose Tissue / Q000166:cytology*; D015152:Blotting, Northern; D002454:Cell Differentiation / Q000235:genetics*; D002469:Cell Separation; D057075:Enzyme Assays; D005786:Gene Expression Regulation; D006801:Humans; D008156:Luciferases / Q000378:metabolism; D059630:Mesenchymal Stem Cells / Q000166:cytology* / Q000378:metabolism*; D035683:MicroRNAs / Q000235:genetics* / Q000378:metabolism; D008967:Molecular Biology / Q000379:methods*; D010012:Osteogenesis / Q000235:genetics*; D020133:Reverse Transcriptase Polymerase Chain Reaction
chemical_list: D035683:MicroRNAs; D008156:Luciferases