Seidel, T.; Artmann, P.J.; Gkekas, I.; Illies, F.; Baack, A.-L.; Viefhues, M. Microfluidic Single-Cell Study on Arabidopsis thaliana Protoplast Fusion—New Insights on Timescales and Reversibilities. Plants2024, 13, 295.
Seidel, T.; Artmann, P.J.; Gkekas, I.; Illies, F.; Baack, A.-L.; Viefhues, M. Microfluidic Single-Cell Study on Arabidopsis thaliana Protoplast Fusion—New Insights on Timescales and Reversibilities. Plants 2024, 13, 295.
Seidel, T.; Artmann, P.J.; Gkekas, I.; Illies, F.; Baack, A.-L.; Viefhues, M. Microfluidic Single-Cell Study on Arabidopsis thaliana Protoplast Fusion—New Insights on Timescales and Reversibilities. Plants2024, 13, 295.
Seidel, T.; Artmann, P.J.; Gkekas, I.; Illies, F.; Baack, A.-L.; Viefhues, M. Microfluidic Single-Cell Study on Arabidopsis thaliana Protoplast Fusion—New Insights on Timescales and Reversibilities. Plants 2024, 13, 295.
Abstract
Plant cells are omnipotent and breeding of new varieties can be achieved by protoplast fusion. Such fusions can be achieved by treatment with poly(ethylene glycol) or by applying an electric field. Microfluidic devices allow for controlled conditions and targeted manipulation of small batches of cells down to single cell analysis. To provide controlled conditions for protoplast fusions and achieve high reproducibility, we developed and characterized a microfluidic device to reliably trap few of Arabidopsis thaliana protoplasts and induced a cell fusion by controlled addition of poly(ethylene glycol) (PEG, with a molecular weight of 6000). Experiments were conducted to determine the survival rate of isolated protoplasts in our microfluidic system. Afterwards, PEG induced fusion was studied. Our results indicate that the following fusion parameters had significant impact on the fusion efficiency and duration: PEG concentration, osmolarity of solution, and flow velocity. PEG concentration below 10% led to only partial fusion. Osmolarity of the PEG fusion solution was found to strongly impact the fusion process; complete fusion of two source cells sufficiently took part in slightly hyper osmotic solutions, whereas iso-osmotic solutions led to only partial fusion at 20% PEG concentration. We observed accelerated fusion for higher fluid velocities. Up to this study, it was common sense that fusion is one directional, i.e. once two cells were fused into one cell they stay fused. Here, we present for the first time reversible fusion of protoplasts. Our microfluidic device paves the way to a deeper understanding on the kinetics and processes of cell fusion.
Copyright:
This is an open access article distributed under the Creative Commons Attribution License which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.
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