Here, we develop and apply a semi-quantitative method for the high-confidence identification of pseudouridylated sites on mammalian mRNAs via direct long-read nanopore sequencing. A comparative analysis of a modification-free transcriptome reveals that the depth of coverage and specific k-mer sequences are critical parameters for accurate basecalling. By adjusting these parameters for high-confidence U-to-C basecalling errors, we identify many known sites of pseudouridylation and uncover previously unreported uridine-modified sites, many of which fall in k-mers that are known targets of pseudouridine synthases. Identified sites are validated using 1000-mer synthetic RNA controls bearing a single pseudouridine in the center position, demonstrating systematic under-calling using our approach. We identify mRNAs with up to 7 unique modification sites. Our workflow allows direct detection of low-, medium-, and high-occupancy pseudouridine modifications on native RNA molecules from nanopore sequencing data and multiple modifications on the same strand.

We report the steady state viscosity and contact microstructure of dense suspensions containing hardparticle poly(methyl methacrylate) (PMMA) colloids with tunable surface morphologies. Structural analysis of confocal micrographs shows that the contact number deficit Δz scales as the jamming distance Δϕ, where the scaling relations contain a range of exponents that describe the compactability of frictional packings with jamming fractions ϕJ and jamming contact numbers zJ. Suspensions with rougher particles require fewer nearest neighbors than that of smoother particles to reach the jamming point. Agreement between model predictions from a mean-field theory and our rheological data shows that shear thickening is modeled by different types of frictional packings that form under applied shear stresses. The shear thickening strength, quantified by the slope of the viscosity-stress flow curves, scales with the jamming distance for a broad class of dense suspensions comprising PMMA smooth and rough colloids, silica smooth and rough colloids, and simulations with interparticle friction or surface asperities. Our results suggest that Δϕ/ϕJ = 0.1 and Δz/zJ = 0.5 is the point at which hydrodynamics, Brownian forces, and friction become equally important in colloidal shear thickening.

Colloidal gels exhibit rich rheological responses under flowing conditions. A clear understanding of the coupling between the kinetics of the formation/rupture of colloidal bonds and the rheological response of attractive gels is lacking. In particular, for gels under different flow regimes, the correlation between the complex rheological response, the bond kinetics, microscopic forces, and an overall micromechanistic view is missing in previous works. Here, we report the bond dynamics in short-range attractive particles, microscopically measured stresses on individual particles and the spatiotemporal evolution of the colloidal structures in different flow regimes. The interplay between interparticle attraction and hydrodynamic stresses is found to be the key to unraveling the physical underpinnings of colloidal gel rheology. Attractive stresses, mostly originating from older bonds dominate the response at low Mason number (the ratio of shearing to attractive forces) while hydrodynamic stresses tend to control the rheology at higher Mason numbers, mostly arising from short-lived bonds. Finally, we present visual mapping of particle bond numbers, their life times and their borne stresses under different flow regimes.