Integration of Thin-Film-Transistor Digital Microfluidics and Cell-Free Systems for the Study of Artificial Cell Compartmentalization via RNA Condensa

Biological systems, while offering a vast library of nano-to-micro engineering solutions, present notable challenges in their understanding due to their inherent complexity. Central to these challenges are experimentation bottleneck presented by the conventional technologies used to interface with them, such as mechanical pipetting or soft-lithography microfluidics. This proposal introduces the development of a platform integrating Thin-Film-Transistor (TFT) Electrowetting-On-Dielectric (EWOD) microfluidics and Cell-Free systems (As Transcription-Translation systems or TX-TL) to create a digitally controlled model of synthetic cells, enabling direct communication between the cells and digital electronics. During this PhD project, we aim to validate the use of EWOD platforms for constructing artificial cells by 1) optimizing the Cell-Free mixtures for EWOD compatibility and 2)characterizing the functionality of the resulting artificial cells by replicating an already established process: the compartmentalization of artificial cells by RNA-based condensates. Once the system is validated, we will adapt existing solutions to construct a platform that allows for further custom hardware and software integration, enabling the expansion of the set of sensors, actuators, and control algorithms. The platform's potential will be demonstrated by exploring the combinatorial space of parameters involved in creating synthetic RNA-condensate organelles, to better understand the impact of different variables on organelle generation, and fine-tuning these parameters to converge into a desired phenotype. By the end of this PhD, we will have developed a prototype of a novel, integrated system that facilitates the bottom-up modeling of cellular processes and apply it to get a better understanding of the compartmentalization processes in artificial cells by programmable RNA condensates.