基于荧光素酶报告基因的细胞微小RNA传感器

Cellular microRNA sensors based on luciferase reporters.

作者信息

Connelly Colleen M, Deiters Alexander

机构信息

Department of Chemistry, North Carolina State University, Raleigh, NC, USA.

出版信息

Methods Mol Biol. 2014;1095:135-46. doi: 10.1007/978-1-62703-703-7_11.

DOI:10.1007/978-1-62703-703-7_11

PMID:24166309

Abstract

Recently, microRNAs (miRNAs) have been linked to a variety of human diseases including cancer and viral infections. Small molecule modifiers of miRNAs could represent new therapeutic agents and be used as tools for elucidating the biological roles of miRNAs. In order to identify small molecule modifiers of miRNAs, functional assays for specific miRNAs must be developed and optimized. Here, we report the construction of a luciferase reporter assay for miRNA miR-122 function and the development of a stable Huh7 cell line that can be used for high-throughput screening of small molecule miR-122 inhibitors. The steps described here can be applied not only to Huh7 cells and miR-122 but also to virtually any cell line and miRNA combination.

摘要

最近，微小RNA（miRNA）已与包括癌症和病毒感染在内的多种人类疾病相关联。miRNA的小分子调节剂可能代表新的治疗剂，并可用作阐明miRNA生物学作用的工具。为了鉴定miRNA的小分子调节剂，必须开发和优化针对特定miRNA的功能测定方法。在此，我们报告了用于miRNA miR-122功能的荧光素酶报告基因测定法的构建，以及一种可用于高通量筛选小分子miR-122抑制剂的稳定Huh7细胞系的开发。这里描述的步骤不仅可以应用于Huh7细胞和miR-122，而且实际上可以应用于任何细胞系和miRNA组合。