DNA Origami Annealing: Folding Yield vs. Cooling Rate

Anneal Program

Cooling rate 0.30 °C/s

Mg²⁺ concentration 12 mM

Start temperature 80 °C

Bath State

Temperature

80.0 °C

Anneal time

0 s

Folding yield

Kinetic traps

Unbound staple site Correctly hybridized Kinetically trapped defect

How it works

Each of ~330 staple-binding sites on the scaffold has its own local melting temperature T_m,i, drawn from a normal distribution (mean set by GC content and Mg²⁺-screened electrostatics, spread ≈ sequence variation across the staple set):

T_m,i ~ N(mu_Tm, sigma_Tm)
mu_Tm(Mg) = 50°C + 6°C * log2(1 + [Mg2+]/6mM)

A site can only find its correct partner while the bath temperature sits inside its narrow hybridization window [T_m,i − w, T_m,i + w] — this is the nucleation-and-zippering step of DNA hybridization. The longer it dwells in that window, the higher its chance of zipping up correctly:

dwell time in window  ∝  1 / cooling_rate
P(correct bind | dt)  =  1 − exp(−k0 · dt)   while T is in window

If the bath drops below a site's window before it has bound — because the cooling rate was too fast — the site freezes into a kinetic trap: a stacked or aggregated misfold that will not anneal out at room temperature. This is exactly why real DNA origami protocols (Rothemund 2006 and successors) ramp slowly, often ~1 hour to overnight from ~80–95°C to room temperature, rather than quenching: slow cooling maximizes the time every staple spends near its own T_m, giving each one a chance to find its correct partner before the structure locks in.

Cooling rate — °C removed from the bath per second of sim time; low = slow careful anneal, high = rapid quench.
Mg²⁺ concentration — divalent cations screen the phosphate backbone's charge, raising every site's T_m and stabilizing correct duplexes (too little Mg²⁺ and staples never bind at all).
Start temperature — the initial melt temperature; must clear every site's T_m + window so the lattice starts fully denatured.

Bath: 80.0 °C

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