Using Genome-Editing Tools to Develop a Novel In Situ Coincidence Reporter Assay for Screening ATAD3A Transcriptional Inhibitors

Liwei Lang, Yong Teng

Research output: Contribution to journalArticle

Abstract

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.

Original languageEnglish (US)
Pages (from-to)159-166
Number of pages8
JournalMethods in molecular biology (Clifton, N.J.)
Volume2138
DOIs
StatePublished - Jan 1 2020

Keywords

  • ATAD3A
  • Coincidence reporter
  • CRISPR-Cas9
  • In situ
  • Transcriptional inhibition

ASJC Scopus subject areas

  • Molecular Biology
  • Genetics

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