Open AccessA Zebrafish Cell Culture Assay for the Identification of MicroRNA Targets
Author(s) -
Weiyi Liu,
Yihong Guan,
Paul Collodi
Publication year - 2010
Publication title -
zebrafish
Language(s) - English
Resource type - Journals
eISSN - 1557-8542
pISSN - 1545-8547
DOI - 10.1089/zeb.2010.0674
Subject(s) - zebrafish , biology , microrna , computational biology , microbiology and biotechnology , hek 293 cells , luciferase , cell culture , regulation of gene expression , genetics , gene , transfection
MicroRNAs (miRNAs) are endogenous small noncoding RNAs that regulate gene expression at the posttranscriptional level. Studies have shown that zebrafish miRNAs play a key role in embryo development, tissue fate establishment, and differentiation by interacting with specific targets, usually in the 3'UTR of the mRNA. Identification of the target sequence is fundamental to elucidating miRNA function. Since bioinformatics can predict hundreds of potential targets for each miRNA, experimental validation of the actual target site is required. Although recent studies have employed the HEK293 cell line to investigate mammalian miRNA targets, our results have shown that the cell line is not suitable for studies of zebrafish miR-430b miRNA. In this article we describe a convenient in vitro assay system that involves the use of zebrafish cell cultures and a luciferase reporter construct to evaluate miR-430b target sites. The cell culture-based assay could be used to validate target sequences of other zebrafish miRNAs.