Version 2 2023-11-20, 16:59Version 2 2023-11-20, 16:59
Version 1 2023-11-20, 05:48Version 1 2023-11-20, 05:48
journal contribution
posted on 2023-11-20, 16:59authored byLien De Wannemaeker, Friederike Mey, Indra Bervoets, Michiel Ver Cruysse, Geoff S. Baldwin, Marjan De Mey
In synthetic biology, Fluorescent reporters are frequently
used
to characterize the expression levels obtained from both genetic parts
such as promoters and ribosome binding sites as well as from complex
genetic circuits. To this end, plate readers offer an easy and high-throughput
way of characterizing both the growth and fluorescence expression
levels of cell cultures. However, despite the similar mode of action
used in different devices, their output is not comparable due to intrinsic
differences in their setup. Additionally, the generated output is
expressed using arbitrary units, limiting reliable comparison of results
to measurements taken within one single experiment using one specific
plate reader, hampering the transferability of data across different
plate readers and laboratories. This article presents an easy and
accessible calibration method for transforming the device-specific
output into a standardized output expressing the amount of fluorescence
per well as a known equivalent fluorophore concentration per cell
for fluorescent reporters spanning the visible light spectrum. This
calibration method follows a 2-fold approach determining both the
estimated number of cells and the equivalent chemical fluorophore
concentration per well. It will contribute to the comparison of plate
reader experiments between different laboratories across the world
and will therefore greatly improve the reliability and exchange of
both results and genetic parts between research groups.