Department of Systems BioMedicine, Tokyo Medical and Dental University (TMDU), Tokyo, Japan.
Department of Molecular Medicine, The Scripps Research Institute, La Jolla, CA, USA.
Methods Mol Biol. 2021;2245:151-166. doi: 10.1007/978-1-0716-1119-7_11.
MicroRNAs (miRNAs) are a class of noncoding small RNAs, which play a critical role in various biological processes including musculoskeletal formation and arthritis pathogenesis via regulating target gene expressions, raising the potentially substantial effects on gene expression networks. Over 2000 miRNAs are encoded in the human genome and a single miRNA potentially targets hundreds of genes. To examine the expression and function of miRNAs in chondrocytes and arthritis pathogenesis, we describe the protocols for the current miRNA related experiments including miRNA expression profiling by (1) Next Generation Sequencing and by TaqMan Array system, (2) miRNA target prediction by TargetScan, (3) miRNA target screening by cell-based reporter library assay, and (4) miRNA and its target interaction by HITS-CLIP (high-throughput sequencing of RNAs isolated by cross-linking immunoprecipitation) in cartilage and chondrocyte research.
MicroRNAs (miRNAs) 是一类非编码的小分子 RNA，通过调节靶基因的表达，在包括肌肉骨骼形成和关节炎发病机制在内的各种生物学过程中发挥着关键作用，对基因表达网络可能产生重大影响。人类基因组中编码了超过 2000 种 miRNAs，一种 miRNA 可能靶向数百个基因。为了研究 miRNA 在软骨细胞和关节炎发病机制中的表达和功能，我们描述了当前 miRNA 相关实验的方案，包括：（1）下一代测序和 TaqMan 微阵列系统的 miRNA 表达谱分析；（2）TargetScan 预测的 miRNA 靶标；（3）基于细胞的报告基因文库测定筛选的 miRNA 靶标；（4）在软骨和软骨细胞研究中通过 HITS-CLIP（交联免疫沉淀分离的 RNA 的高通量测序）分析 miRNA 及其靶标相互作用。
