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The dual luciferase reporter system and RT‐qPCR strategies for screening of MicroRNA‐21 small‐molecule inhibitors
Author(s) -
Hei YuanYuan,
Guo YuanXu,
Jiang CongShan,
Wang Si,
Lu SheMin,
Zhang SanQi
Publication year - 2019
Publication title -
biotechnology and applied biochemistry
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.468
H-Index - 70
eISSN - 1470-8744
pISSN - 0885-4513
DOI - 10.1002/bab.1756
Subject(s) - luciferase , small molecule , enoxacin , microrna , chemistry , reporter gene , norfloxacin , plasmid , computational biology , biology , combinatorial chemistry , biochemistry , transfection , gene expression , gene , antibiotics , ciprofloxacin
Abstract The therapeutic potential of microRNA‐21 (miR‐21) small‐molecule inhibitors has been of particular interest to medicinal chemists. Moreover, the development of more facile screening methods is lacking. In the present study, two potential screening strategies for miR‐21 small‐molecule inhibitor including the stem‐loop reverse transcription‐quantitative PCR and dual luciferase reporter assay system were demonstrated and discussed in detail. A pmirGLO‐miR21cswt plasmid and its two different mutants were constructed for dual luciferase reporter assay system. In addition, the sensitivity and specificity of these two methods were validated. Our results demonstrated that both strategies are decent choices for the screening of small‐molecule inhibitors for miR‐21 and possibly other miRNAs. Eventually, we applied our optimized strategy to discover and characterize several promising compounds such as azobenzene derivate A, enoxacin, and norfloxacin for their potential impact on intracellular miR‐21 concentration.