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Abstract Many viruses eject their DNA via a nanochannel in the viral shell, driven by internal forces arising from the high-density genome packing. The speed of DNA exit is controlled by friction forces that limit the molecular mobility, but the nature of this friction is unknown. We introduce a method to probe the mobility of the tightly confined DNA by measuring DNA exit from phage phi29 capsids with optical tweezers. We measure extremely low initial exit velocity, a regime of exponentially increasing velocity, stochastic pausing that dominates the kinetics and large dynamic heterogeneity. Measurements with variable applied force provide evidence that the initial velocity is controlled by DNA–DNA sliding friction, consistent with a Frenkel–Kontorova model for nanoscale friction. We confirm several aspects of the ejection dynamics predicted by theoretical models. Features of the pausing suggest that it is connected to the phenomenon of ‘clogging’ in soft matter systems. Our results provide evidence that DNA–DNA friction and clogging control the DNA exit dynamics, but that this friction does not significantly affect DNA packaging. 

We previously introduced the use of DNA molecules for calibration of biophysical force and displacement measurements with optical tweezers. Force and length scale factors can be determined from measurements of DNA stretching. Trap compliance can be determined by fitting the data to a nonlinear DNA elasticity model, however, noise/drift/offsets in the measurement can affect the reliability of this determination. Here we demonstrate a more robust method that uses a linear approximation for DNA elasticity applied to high force range (25–45 pN) data. We show that this method can be used to assess how small variations in microsphere sizes affect DNA length measurements and demonstrate methods for correcting for these errors. We further show that these measurements can be used to check assumed linearities of system responses. Finally, we demonstrate methods combining microsphere imaging and DNA stretching to check the compliance and positioning of individual traps.