Title

Thermodynamics Of DNA Duplex Formation: A Biophysical Chemistry Laboratory Experiment

Document Type

Article

Publication Date

11-1-2000

Published In

Journal Of Chemical Education

Abstract

The goal of this biophysical experiment is to measure thermodynamic properties of a short DNA duplex by melting the ordered native structure (duplex) into the disordered, denatured state (single strands) while monitoring the transition using ultraviolet spectrophotometry. The DNA samples we used were two complementary synthetic DNA oligomers, dCA7G and dCT7G. The absorbance of four separate DNA samples with different concentrations (10-75 μM for each single strand) is measured at 260 nm as the temperature is increased from 10 to 70 °C. As the ordered regions of stacked base pairs in the DNA duplex are disrupted, the UV absorbance increases. The profile of UV absorbance versus temperature is called a melting curve and the midpoint of the transition is defined as the melting temperature, Tm. The dependence of Tm on strand concentration can be analyzed to yield quantitative thermodynamic data (ΔH°, ΔS°, ΔG°).

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