= RNA melting hysteresis
{c}
<RNA melting hysteresis> is dependence of an <RNA optical melting> trace on thermal history: heating and cooling can give different <absorbance> at the same <temperature>. It warns that the observed trace may not follow reversible <chemical equilibrium> on the measurement timescale. Two different traces do not by themselves establish two equilibrium melting temperatures.
A primary experiment illustrates the importance of history. A 21-nucleotide <RNA> showed a low-temperature dimer-to-hairpin transition that did not reverse on cooling, whereas its higher-temperature <RNA hairpin melting> transition was reversible. Measurements used a Cary 100 <spectrophotometer>, a 1-cm quartz cell and 268-nm light, heating at $1.0\,{}^{\circ}\mathrm C/\mathrm{min}$ and cooling at $0.5$ or $0.2\,{}^{\circ}\mathrm C/\mathrm{min}$. An adjacent solvent cell held the temperature probe.<Conversion of stable RNA hairpin to a metastable dimer in frozen solution by Xueguang Sun and others>[Results, Fig.5 and transition reversibility; Materials and Methods, UV absorbance melting curves.]{ref} The authors interpreted the dimer as kinetically trapped under those conditions, rather than applying a reversible equilibrium fit to that transition.
For an original simple kinetic model, let $f$ be the unfolded fraction, $f_{\rm eq}(T)$ its equilibrium value and $\tau$ an <RNA melting relaxation time>:
$$
\frac{df}{dt}=\frac{f_{\rm eq}(T)-f}{\tau}.
$$
After a fixed-temperature step with constant $\tau$, the deviation undergoes <exponential decay>:
$$
f(t)=f_{\rm eq}+(f(0)-f_{\rm eq})e^{-t/\tau}.
$$
With stipulated $f(0)=0$, $f_{\rm eq}=1$ and $\tau=2\,\mathrm{min}$, the fraction reaches only 0.632 after two minutes and 0.950 after six. During a continuing temperature ramp, this lag can put heating and cooling on different branches. This single-time model is illustrative; it is not a fit to the published dimer experiment.
Useful checks compare ramp speeds and include fixed-temperature holds and repeat cycles. Verify <measurement calibration> and exclude evaporation or chemical degradation before attributing a discrepancy to folding kinetics. Slower scans can reduce kinetic lag, but a smooth trace alone cannot establish reversibility.
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