Department of Bioinformatics and Genomics, The University of North Carolina at Charlotte, Charlotte, North Carolina, USA.
PLoS One. 2010 Jun 10;5(6):e11048. doi: 10.1371/journal.pone.0011048.
The probe percent bound value, calculated using multi-state equilibrium models of solution hybridization, is shown to be useful in understanding the hybridization behavior of microarray probes having 50 nucleotides, with and without mismatches. These longer oligonucleotides are in widespread use on microarrays, but there are few controlled studies of their interactions with mismatched targets compared to 25-mer based platforms.
50-mer oligonucleotides with centrally placed single, double and triple mismatches were spotted on an array. Over a range of target concentrations it was possible to discriminate binding to perfect matches and mismatches, and the type of mismatch could be predicted accurately in the concentration midrange (100 pM to 200 pM) using solution hybridization modeling methods. These results have implications for microarray design, optimization and analysis methods.
Our results highlight the importance of incorporating biophysical factors in both the design and the analysis of microarrays. Use of the probe "percent bound" value predicted by equilibrium models of hybridization is confirmed to be important for predicting and interpreting the behavior of long oligonucleotide arrays, as has been shown for short oligonucleotide arrays.
使用溶液杂交平衡模型计算的探针结合百分比值，被证明有助于理解具有 50 个核苷酸的微阵列探针的杂交行为，无论是否存在错配。这些较长的寡核苷酸在微阵列中被广泛使用，但与基于 25 -mer 的平台相比，对它们与错配靶标相互作用的研究很少。
在阵列上点样了中央有单个、双个和三个错配的 50-mer 寡核苷酸。在一系列靶标浓度范围内，可以区分与完美匹配和错配的结合，并且可以使用溶液杂交建模方法在浓度中程（100 pM 至 200 pM）准确预测错配的类型。这些结果对微阵列设计、优化和分析方法有影响。
我们的结果强调了在微阵列的设计和分析中纳入生物物理因素的重要性。使用杂交平衡模型预测的探针“结合百分比”值对于预测和解释长寡核苷酸阵列的行为非常重要，正如短寡核苷酸阵列所示。
