PlotXpress, a webtool for normalization and visualization of reporter expression data

Elias Brandorff,Joachim Goedhart,Marc Galland

doi:10.12688/f1000research.73641.1

Abstract

In molecular cell biology, reporter assays are frequently used to investigate gene expression levels. Reporter assays employ a gene that encodes a light-emitting protein, of which the luminescence is quantified as a proxy of gene expression. Commercial parties provide reporter assay kits that include protocols and specialized detection machinery. However, downstream analysis of the output data and their presentation are not standardized. We have developed plotXpress to fill this gap, providing a free, open-source platform for the semi-automated analysis and standardized visualisation of experimental gene reporter data. Users can upload raw luminescence data acquired from a reporter gene assay with an internal control. In plotXpress, the data is corrected for sample variation with the internal control and the average for each condition is calculated. When a reference condition is selected the fold change is calculated for all other conditions, based on the selected reference. The results are shown as dot plots with a statistical summary, which can be adjusted to create publication-grade plots without requiring coding skills. Altogether, plotXpress is an open-source, low-threshold, web-based tool, that promotes a standardized and reproducible analysis while providing an appealing visualization of reporter data.The webtool can be accessed at: https://huygens.science.uva.nl/PlotXpress/.

Highlights

Reporter gene assays are popular tools to investigate gene expression dynamics in cell biology (Barriscale et al, 2014; Liu et al, 2009; Schenborn and Groskreutz, 1999)
The expression level of the reporter gene is used as a proxy of the capacity of a DNA sequence to regulate gene expression under various experimental conditions
Reporter gene activity can be compared between variations of a regulatory DNA sequence, for example by removing a transcription factor binding site or by modulating tandem repeat expansions in a promoter sequence (Rodriguez et al, 2020; Moparthi and Koch, 2020)

Summary

Introduction

Reporter gene assays are popular tools to investigate gene expression dynamics in cell biology (Barriscale et al, 2014; Liu et al, 2009; Schenborn and Groskreutz, 1999). Reporter gene activity can be compared between variations of a regulatory DNA sequence, for example by removing a transcription factor binding site or by modulating tandem repeat expansions in a promoter sequence (Rodriguez et al, 2020; Moparthi and Koch, 2020). In the so-called “Dual-LuciferaseR Reporter Assay System” (Promega), an internal control is used to correct for variations in cell density and transfection efficiency. This internal control is co-transfected and consists of a vector with a constitutively active promoter that drives expression of another luciferase. As coding skills are not required, plotXpress is a readily available low entry-level application that democratizes the processing and visualization of dual reporter expression data

Methods

Conclusion