Dorris David R, Nguyen Allen, Gieser Linn, Lockner Randall, Lublinsky Anna, Patterson Marcus, Touma Edward, Sendera Timothy J, Elghanian Robert, Mazumder Abhijit
Motorola Life Sciences, Northbrook, IL 60062, USA.
BMC Biotechnol. 2003 Jun 11;3:6. doi: 10.1186/1472-6750-3-6.
DNA microarrays are now routinely used to monitor the transcript levels of thousands of genes simultaneously. However, the array fabrication method, hybridization conditions, and oligodeoxyribonucleotide probe length can impact the performance of a DNA microarray platform.
We demonstrate solution-phase hybridization behavior of probe:target interactions by showing a strong correlation between the effect of mismatches in probes attached to a three dimensional matrix of a microarray and solution-based, thermodynamic duplex melting studies. The effects of mismatches in the probes attached to the microarray also demonstrate that most, if not all, of the oligodeoxyribonucleotide is available for hybridization. Kinetic parameters were also investigated. As anticipated, hybridization signals increased in a transcript concentration-dependent manner, and mismatch specificity increased with hybridization time. Unexpectedly, hybridization time increased the accuracy of fold changes by relieving the compression observed in expression ratios, and this effect may be more dramatic for larger fold changes.
Taken together, these studies demonstrate that a three-dimensional surface may enable use of shorter oligodeoxyribonucleotide probes and that hybridization time may be critical in improving the accuracy of microarray data.
DNA微阵列目前常用于同时监测数千个基因的转录水平。然而，阵列制造方法、杂交条件和寡脱氧核糖核苷酸探针长度会影响DNA微阵列平台的性能。
我们通过展示附着在微阵列三维基质上的探针中错配效应与基于溶液的热力学双链解链研究之间的强相关性，证明了探针与靶标相互作用的溶液相杂交行为。附着在微阵列上的探针中错配的影响还表明，大多数（如果不是全部）寡脱氧核糖核苷酸可用于杂交。还研究了动力学参数。正如预期的那样，杂交信号以转录本浓度依赖的方式增加，错配特异性随杂交时间增加。出乎意料的是，杂交时间通过缓解表达比率中观察到的压缩来提高倍数变化的准确性，并且对于更大的倍数变化，这种效应可能更显著。
综上所述，这些研究表明三维表面可能允许使用更短的寡脱氧核糖核苷酸探针，并且杂交时间对于提高微阵列数据的准确性可能至关重要。
