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DNA multiplex hybridization on microarrays and thermodynamic stability in solution: a direct comparison.

Fish DJ, Horne MT, Brewood GP, Goodarzi JP, Alemayehu S, Bhandiwad A, Searles RP, Benight AS - Nucleic Acids Res. (2007)

Bottom Line: Hybridization intensities of 30 distinct short duplex DNAs measured on spotted microarrays, were directly compared with thermodynamic stabilities measured in solution.Quantitative comparison with results from 63 multiplex microarray hybridization experiments provided a linear relationship for perfect match and most mismatch duplexes.These observations underscore the need for rigorous evaluation of thermodynamic parameters describing tandem mismatch stability.

View Article: PubMed Central - PubMed

Affiliation: Portland Bioscience, Inc., Portland State University, USA. djf@pdxbio.com

ABSTRACT
Hybridization intensities of 30 distinct short duplex DNAs measured on spotted microarrays, were directly compared with thermodynamic stabilities measured in solution. DNA sequences were designed to promote formation of perfect match, or hybrid duplexes containing tandem mismatches. Thermodynamic parameters DeltaH degrees , DeltaS degrees and DeltaG degrees of melting transitions in solution were evaluated directly using differential scanning calorimetry. Quantitative comparison with results from 63 multiplex microarray hybridization experiments provided a linear relationship for perfect match and most mismatch duplexes. Examination of outliers suggests that both duplex length and relative position of tandem mismatches could be important factors contributing to observed deviations from linearity. A detailed comparison of measured thermodynamic parameters with those calculated using the nearest-neighbor model was performed. Analysis revealed the nearest-neighbor model generally predicts mismatch duplexes to be less stable than experimentally observed. Results also show the relative stability of a tandem mismatch is highly dependent on the identity of the flanking Watson-Crick (w/c) base pairs. Thus, specifying the stability contribution of a tandem mismatch requires consideration of the sequence identity of at least four base pair units (tandem mismatch and flanking w/c base pairs). These observations underscore the need for rigorous evaluation of thermodynamic parameters describing tandem mismatch stability.

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Difference in change in enthalpy, ΔΔHend − ΔΔHmid (dark) and change in entropy, TΔΔSend − TΔΔSmid (light), for moving a tandem mismatch from the middle to near the end of the duplex, i.e. from Type 2 duplex (mismatch in the middle: 6.2, 7.2 or 10.2) to Type 3 duplex (mismatch near the end: 1.3 or 3.3).
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Figure 9: Difference in change in enthalpy, ΔΔHend − ΔΔHmid (dark) and change in entropy, TΔΔSend − TΔΔSmid (light), for moving a tandem mismatch from the middle to near the end of the duplex, i.e. from Type 2 duplex (mismatch in the middle: 6.2, 7.2 or 10.2) to Type 3 duplex (mismatch near the end: 1.3 or 3.3).

Mentions: In order to emphasize these results, the differences between ΔΔH (and TΔΔS) for these duplexes are shown in Figure 9. These values represent the effective difference in ΔH° and TΔS° upon replacing a tandem mismatch in the middle of the duplex with one near the end. As expected from the data in Figure 8, the differences in ΔΔH (and TΔΔS) decrease with decreasing stability of the flanking base pairs. The relative cost of replacing a tandem mismatch in the middle with one on the end (6.2, 7.2, 10.2 → 1.3) follows the following order: ΔΔH(1.3) − ΔΔH(7.2) > ΔΔH(1.3) − ΔΔH(6.2) > ΔΔH(1.3) − ΔΔH(10.2) (similarly for TΔΔS). The relative cost of replacing a tandem mismatch in the middle with one on the penultimate end (6.2, 7.2, 10.2 → 3.3) also follows the same order. In both cases, the relative order of the differences between mismatches in the middle and on the ends is maintained. This observation indicates that the sequence dependent thermodynamics of flanking w/c base pairs have a greater influence than mismatch position on tandem mismatch stability.Figure 9.


DNA multiplex hybridization on microarrays and thermodynamic stability in solution: a direct comparison.

Fish DJ, Horne MT, Brewood GP, Goodarzi JP, Alemayehu S, Bhandiwad A, Searles RP, Benight AS - Nucleic Acids Res. (2007)

Difference in change in enthalpy, ΔΔHend − ΔΔHmid (dark) and change in entropy, TΔΔSend − TΔΔSmid (light), for moving a tandem mismatch from the middle to near the end of the duplex, i.e. from Type 2 duplex (mismatch in the middle: 6.2, 7.2 or 10.2) to Type 3 duplex (mismatch near the end: 1.3 or 3.3).
© Copyright Policy - creative-commons
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC2175334&req=5

Figure 9: Difference in change in enthalpy, ΔΔHend − ΔΔHmid (dark) and change in entropy, TΔΔSend − TΔΔSmid (light), for moving a tandem mismatch from the middle to near the end of the duplex, i.e. from Type 2 duplex (mismatch in the middle: 6.2, 7.2 or 10.2) to Type 3 duplex (mismatch near the end: 1.3 or 3.3).
Mentions: In order to emphasize these results, the differences between ΔΔH (and TΔΔS) for these duplexes are shown in Figure 9. These values represent the effective difference in ΔH° and TΔS° upon replacing a tandem mismatch in the middle of the duplex with one near the end. As expected from the data in Figure 8, the differences in ΔΔH (and TΔΔS) decrease with decreasing stability of the flanking base pairs. The relative cost of replacing a tandem mismatch in the middle with one on the end (6.2, 7.2, 10.2 → 1.3) follows the following order: ΔΔH(1.3) − ΔΔH(7.2) > ΔΔH(1.3) − ΔΔH(6.2) > ΔΔH(1.3) − ΔΔH(10.2) (similarly for TΔΔS). The relative cost of replacing a tandem mismatch in the middle with one on the penultimate end (6.2, 7.2, 10.2 → 3.3) also follows the same order. In both cases, the relative order of the differences between mismatches in the middle and on the ends is maintained. This observation indicates that the sequence dependent thermodynamics of flanking w/c base pairs have a greater influence than mismatch position on tandem mismatch stability.Figure 9.

Bottom Line: Hybridization intensities of 30 distinct short duplex DNAs measured on spotted microarrays, were directly compared with thermodynamic stabilities measured in solution.Quantitative comparison with results from 63 multiplex microarray hybridization experiments provided a linear relationship for perfect match and most mismatch duplexes.These observations underscore the need for rigorous evaluation of thermodynamic parameters describing tandem mismatch stability.

View Article: PubMed Central - PubMed

Affiliation: Portland Bioscience, Inc., Portland State University, USA. djf@pdxbio.com

ABSTRACT
Hybridization intensities of 30 distinct short duplex DNAs measured on spotted microarrays, were directly compared with thermodynamic stabilities measured in solution. DNA sequences were designed to promote formation of perfect match, or hybrid duplexes containing tandem mismatches. Thermodynamic parameters DeltaH degrees , DeltaS degrees and DeltaG degrees of melting transitions in solution were evaluated directly using differential scanning calorimetry. Quantitative comparison with results from 63 multiplex microarray hybridization experiments provided a linear relationship for perfect match and most mismatch duplexes. Examination of outliers suggests that both duplex length and relative position of tandem mismatches could be important factors contributing to observed deviations from linearity. A detailed comparison of measured thermodynamic parameters with those calculated using the nearest-neighbor model was performed. Analysis revealed the nearest-neighbor model generally predicts mismatch duplexes to be less stable than experimentally observed. Results also show the relative stability of a tandem mismatch is highly dependent on the identity of the flanking Watson-Crick (w/c) base pairs. Thus, specifying the stability contribution of a tandem mismatch requires consideration of the sequence identity of at least four base pair units (tandem mismatch and flanking w/c base pairs). These observations underscore the need for rigorous evaluation of thermodynamic parameters describing tandem mismatch stability.

Show MeSH