Using Genome-Editing Tools to Develop a Novel In Situ Coincidence Reporter Assay for Screening ATAD3A Transcriptional Inhibitors
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Transgene-based reporter gene assays have been used for discovery of inhibitors targeting vital gene transcription. In traditional assays, the reporter gene is commonly fused with a cloned promoter and integrated into a random genomic location. This has been widely applied but significantly dampened by disadvantages, including incomplete cis-acting elements, the influence of foreign epigenetic environments, and generation of false hits that disrupt the luciferase reporter activity. Therefore, there is a need to develop novel strategies for developing in situ reporter assays closely mimicking endogenous gene expression without disrupting its function. By employing the CRISPR-Cas9 system, we developed an effective in situ coincidence reporter system with a selection marker in the endogenous locus of ATAD3A, which provides a means of screening for transcription-targeted lead compounds with high confidence.
Key wordsATAD3A CRISPR-Cas9 Coincidence reporter In situ Transcriptional inhibition
This research was supported by NIH grant R03DE028387 and R01DE028351 (to Y.T.).
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