Fig. 4: Sequence-orthogonal RNA nanostars produce distinct condensates that recruit clients specifically and remain demixed.

a Multi-stranded sequence-orthogonal nanostars modified to include FLAPs. When annealed in one pot they produce distinct RNA condensates that do not mix. Condensates were imaged in both FITC and Cy3 channels; each pixel’s FITC/Cy3 ratio was mapped to an angle θ and histogrammed. Histogram peaks for nanostars annealed individually align with those of nanostars annealed together. b Top: Nanostars including aptamer domains that recruit streptavidin, P22 N peptide, and TAT peptide guests. Middle: Example images and violin plots of the CLD for condensates before and after recruitment of peptides. Black line indicates the mean. Bottom: Nanostars were designed to be sequence-orthogonal, thus their one-pot assembly with fluorescently labeled clients shows their recruitment and compartmentalized into specific condensates (red, blue, and yellow). c Sequence-orthogonal single-stranded RNA motifs pairs (two nanostar systems, 3sβ and 3sγ), modified to include Corn and Red Broccoli fluorogenic aptamers. The peaks of the FITC/Cy3 angle histogram for nanostars annealed individually align with the peaks of nanostars annealed together. Experiments were done in triplicates; images provide representative examples. Violin plots and histograms pool data from all replicates. Scale bars: 20 µm.