Dresden 2017 – wissenschaftliches Programm

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BP: Fachverband Biologische Physik

BP 33: Posters - Systems Biology & Gene Expression and Signalling

BP 33.5: Poster

Dienstag, 21. März 2017, 14:00–16:00, P2-OG1

Deconvolution of luminescence cross-talk in high-throughput gene expression profiling — •Marco Mauri, Stefano Vecchione, and Georg Fritz — LOEWE-Center for Synthetic Microbiology (SYNMIKRO), Philipps-University Marburg, Germany

In recent years, luciferase has become a standard genetic tool to monitor gene expression. It has a high signal-to-noise ratio, which is, in principle, only limited by the sensitivity of the photo detector. However, at the same time luciferase reporters have the drawback of emitting a constant glow upon induction, which can lead to undesired cross-talk between neighbouring wells on a microplate. Indeed, we find that the scattering light from a highly luminescent well affects more than 50% of the wells even in a black plate. In order to overcome this limitation, we developed a computational method to correct for luminescence bleed-through and estimate the "true" luminescence activity for each well of a microplate. As the sole input to our algorithm the signals measured from a calibration plate is needed, in which the light emitted from a single luminescent well serves as an estimate of the light-spread function. From this the algorithm creates a deconvolution matrix, which can be used to correct any other measurement obtained under the same technical conditions. Here, we demonstrate that our correction preserves low level signals that are close to the background and show that it is universally applicable to different kinds of microplate readers and plate types. From our algorithm, we developed a freely available tool to correct the luminescence cross-talk in high-throughput gene expression analyses.