Format

Send to

Choose Destination
Methods Mol Biol. 2018;1755:163-177. doi: 10.1007/978-1-4939-7724-6_12.

Endogenous Locus Reporter Assays.

Author information

1
Screening & Protein Sciences, Merck Research Labs, Merck & Co., Inc., West Point, PA, USA.
2
Screening and Translational Enzymology, Roche, Basel, Roche, Basel, Canton of Basel-Stadt, Switzerland.
3
Screening & Protein Sciences, Merck Research Labs, Merck & Co., Inc., West Point, PA, USA. matthew_tudor@merck.com.

Abstract

Reporter gene assays are widely used in high-throughput screening (HTS) to identify compounds that modulate gene expression. Traditionally a reporter gene assay is built by cloning an endogenous promoter sequence or synthetic response elements in the regulatory region of a reporter gene to monitor transcriptional activity of a specific biological process (exogenous reporter assay). In contrast, an endogenous locus reporter has a reporter gene inserted in the endogenous gene locus that allows the reporter gene to be expressed under the control of the same regulatory elements as the endogenous gene, thus more accurately reflecting the changes seen in the regulation of the actual gene. In this chapter, we introduce some of the considerations behind building a reporter gene assay for high-throughput compound screening and describe the methods we have utilized to establish 1536-well format endogenous locus reporter and exogenous reporter assays for the screening of compounds that modulate Myc pathway activity.

KEYWORDS:

Endogenous locus reporter; Exogenous reporter; High-throughput screening (HTS); Luciferase; Myc; NanoLuc; PEST; Reporter gene

PMID:
29671270
DOI:
10.1007/978-1-4939-7724-6_12
[Indexed for MEDLINE]

Supplemental Content

Full text links

Icon for Springer
Loading ...
Support Center