Project Summary: Administrative Supplement to NIGMS MIRA R35GM122561 Parent project title: Dynamics and evolution of synthetic and natural gene regulatory networks This Administrative Supplement is based on NOT-GM-21-030, “Notice of Special Interest (NOSI): Administrative Supplements for Equipment Purchases for NIGMS R01, R35, and R37 Awardees”. It is a request for partial support of a FluidFM Bio system by Cytosurge AG, a Swiss Biotech company. The FluidFM system has unique capabilities for repeated, single live cell-specific, direct intra- cytoplasmic or intra-nuclear delivery and extraction of fluids, as well as nondestructive single cell manipulation. These capabilities will be transformative in promoting progress with the parent grant, which aims to learn how gene network dynamics and stochasticity affect single cells and thereby cell populations. To achieve this, we proposed using synthetic gene networks to generate specific gene expression patterns in space and time that serve as signals for natural gene networks, studying the subsequent effects on cell population behavior and evolution by computational modeling and experimental evolution. The FluidFM system will accelerate the stable genomic insertion of synthetic gene circuits by coupled intranuclear delivery of CRISPR, recombinase and genetic payloads into single cells, by cell isolation for clonal outgrowth, and by repeated “biopsies” taken from engineered single living cells to study how their transcriptomic and proteomic networks respond to perturbations by synthetic gene circuits. Overall, the FluidFM system will provide unique capabilities to illuminate how complex networks enable control across scales of space and time in biology, from molecules to cells. Addressing these questions will teach us how to control adapting cell populations, which is relevant for understanding, predicting and possibly preventing cancer and microbial drug resistance.