Cell - 8 September 2016

Figure S1. Characterization of PGL-3 Drop Properties in the Presence or Absence of mRNA, Related toFigures 1and 3
(A and B) Maximum intensity projections of series of confocal z- slices show different concentrations of PGL-3-mEGFP in absence (A) or in presence of 50 ng/mL
(or 100 nM) mouse brain mRNA (B).
(C) Plot of total GFP fluorescence found in drops and surrounding bulk phase as a function of PGL-3-mEGFP concentration in presence or absence of 50 ng/mL (or
100 nM) mouse brain mRNA. For each concentration of PGL-3-mEGFP, drops inR 12 observation volumes (41mm 341 mm 3  10 mm) were scored. Error bars
represent 1 SEM among the observation volumes scored. Blue circles: no RNA, red circles: in presence of mRNA. Blue and red lines represent linear fits (R^2 = 0.9
or 0.75).
(D) Bright-field image shows PGL-3 at 10mM phase-separates into drops in absence of any polypeptide tags.
(E) PGL-3-mEGFP forms drops after the protein has been treated with the nuclease Benzonase. PGL-3-mEGFP at 0.5mM was incubated with 125 units of
Benzonase for 4 hr at room temperature. Maximum intensity projection of series of confocal z- slices shows PGL-3-mEGFP retained the ability to phase-separate
into drops once the dilute protein solution was concentrated following treatment with Benzonase.
(F) Measurement of the material properties of drops of full-length PGL-3 in controlled fusion experiments conducted in a dual-trap optical tweezer microscope.
Plot of fusion time as a function of size of droplets undergoing fusion. The data were fitted with a straight line the slope of which provides the ratio of surface
tension and viscosity (h/n).
(G) Time-lapse single confocal plane micrographs show fluorescence recovery of PGL-3-mEGFP (at 10mM) after a full PGL-3-mEGFP drop is photobleached
at 6 s.
(H) Time-lapse single confocal plane micrographs show recovery of fluorescence following photobleaching a PGL-3-mEGFP drop (at 0.6mM) assembled on
addition of 10 ng/mL unlabeled totalC. elegansmRNA.
(I) FRAP data to measure the exchange of PGL-3 between drop and surroundings in presence or absence of RNA. Black trace: photobleaching of 21 drops of
PGL-3-mEGFP at 10mM in absence of RNA, Red trace: photobleaching of 24 drops of PGL-3-mEGFP at 0.6mM assembled on addition of 10 ng/mL total
C. elegansmRNA. Error bars represent 1 SD.