Full details of all procedures, including specific compounds used, specific details surrounding phase separation assays, biolayer interferometry, FRAP assays, droplet fusion experiments, partitioning assays, SXT, and coarse-grained simulations are provided in SI Appendix .
For phase separation assays, polyanions were diluted to 1 µg/µL in 100 mM K2HPO4/KH2PO4 buffer at pH 7. Peptides were added at 250-µM concentrations. For partitioning experiments, probe molecules were spiked in at 100 nM. For the generation of pure RNA liquid droplets, homopolymeric RNAs were diluted to 2 µg/µL in 1x PBS buffer with 30% PEG and 10 mM MgCl2 at pH 7. For intradroplet FRAP, a circular area of 1-μM radius was bleached in droplets with a radius between 5 μM and 10 μM. For SXT 3D reconstructions, 92 projection images, with 200-ms exposure time each, were acquired sequentially around a rotation axis with 2° increment angles. For droplet fusion assays, confocal time-lapse images were taken of droplets settling on the coverslip and fusing together. Coarse-grained simulations are lattice-based and utilize a simple neighbor−neighbor interaction potential. For all other details, please seeSI Appendix .
For phase separation assays, polyanions were diluted to 1 µg/µL in 100 mM K2HPO4/KH2PO4 buffer at pH 7. Peptides were added at 250-µM concentrations. For partitioning experiments, probe molecules were spiked in at 100 nM. For the generation of pure RNA liquid droplets, homopolymeric RNAs were diluted to 2 µg/µL in 1x PBS buffer with 30% PEG and 10 mM MgCl2 at pH 7. For intradroplet FRAP, a circular area of 1-μM radius was bleached in droplets with a radius between 5 μM and 10 μM. For SXT 3D reconstructions, 92 projection images, with 200-ms exposure time each, were acquired sequentially around a rotation axis with 2° increment angles. For droplet fusion assays, confocal time-lapse images were taken of droplets settling on the coverslip and fusing together. Coarse-grained simulations are lattice-based and utilize a simple neighbor−neighbor interaction potential. For all other details, please see
