Melting of a DNA Triplex without Hyperchromism.

 

Fu-Sung Hung1,2, Tsung-Mei Chin1, and Lou-Sing Kan2

1Institute of Applied Chemistry, Chinese Culture University, 2Institute of Chemistry, Academia Sinica, Taipei, Taiwan

 

UV absorbance spectroscopy is the most common method for detecting nucleic acid structural transitions and obtaining thermodynamic parameters.  We report that in some cases UV absorbance spectroscopy is an inadequate analytical technique for these purposes.  Some critical transitions are invisible to UV absorbance or its derivative with temperature.  The Hoogsteen strand dissociation of a hairpin type triplex, composed of the probe 5'-d-(AG)nT4(CT)n and the target 5'-d-(TC)n/5'-d-(CT)n (n = 3-5), is not accompanied by hyperchromism at 260 nm, although there is a significant change in the extent of base stacking.  The UV-invisible transition we describe here is observed by the ethidium bromide fluorescence enhancement study.  The triplex formation is further confirmed by comparison to the CD spectra.  The results show that structural transitions may occur without producing the expected signals.  Our observation challenges the use of hyperchromicity for monitoring nucleic acid structural transitions.

 

 

 

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