DNA multiplex hybridization on microarrays and thermodynamic stability in solution: a direct comparison.
DNA multiplex hybridization on microarrays and thermodynamic stability in solution: a direct comparison.
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在微阵列上的DNA多重杂交和溶液中的热力学稳定性：直接比较。
DOI：
10.1093/nar/gkm865
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发表时间：
2007
影响因子：
14.9
通讯作者：
Benight, Albert S
中科院分区：
文献类型：
--
作者：
Fish, Daniel J;Horne, M Todd;Brewood, Greg P;Goodarzi, Jim P;Alemayehu, Saba;Bhandiwad, Ashwini;Searles, Robert P;Benight, Albert S
在斑点微阵列上测量的30种不同的短双链DNA的杂交强度与在溶液中测量的热力学稳定性直接进行比较。设计DNA序列以促进完全匹配或含有串联错配的杂交双链体的形成。用差示扫描量热法直接测定了溶液熔融转变的热力学参数ΔH°、ΔS°和ΔG°。与63个多重微阵列杂交实验的结果定量比较，为完全匹配和大多数错配双链体提供了线性关系。离群值检查表明，双链体长度和串联错配的相对位置可能是导致观察到的线性偏差的重要因素。测量的热力学参数与使用最近邻模型计算的热力学参数进行了详细的比较。分析显示，最近邻模型通常预测错配双链体比实验观察到的更不稳定。结果还表明，串联错配的相对稳定性高度依赖于侧翼沃森-克里克（w/c）碱基对的身份。因此，指定串联错配的稳定性贡献需要考虑至少四个碱基对单元（串联错配和侧翼w/c碱基对）的序列同一性。这些意见强调需要严格的热力学参数评价描述串联失配稳定性。
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 ΔH°, ΔS° and ΔG° 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.