FlopR: An Open Source Software Package for Calibration and Normalization of Plate Reader and Flow Cytometry Data

Area of Science:

• Synthetic biology
• Quantitative biology
• Microbiology

Background:

• Gene expression measurement using fluorescence is crucial in synthetic biology.
• Arbitrary units in fluorescence data limit quantitative applications.
• Existing calibration tools for flow cytometry and plate readers are fragmented.

Purpose of the Study:

• To develop a unified software tool for calibrating fluorescence measurements.
• To improve the quantitative utility of gene expression data in synthetic biology.
• To enhance data interoperability within the synthetic biology community.

Main Methods:

• Development of an R package integrating and improving existing calibration methods.
• Validation using *Escherichia coli* engineered to express green fluorescent protein (GFP).
• Application of the tool to analyze time-evolution of bacterial subpopulations from plate reader data.

Main Results:

• A single, cohesive software tool for fluorescence measurement calibration is presented.
• The R package successfully calibrates fluorescence data from engineered *E. coli*.
• The tool enables identification of bacterial subpopulations from bulk plate reader data, reducing reliance on flow cytometry.

Conclusions:

• The developed R package provides a standardized approach to quantitative fluorescence measurements.
• This tool enhances data analysis capabilities in synthetic biology, particularly for bacterial subpopulation dynamics.
• Standardized tools are essential for advancing interoperability and quantitative rigor in synthetic biology research.