Absolute mRNA concentrations from sequence-specific calibration of oligonucleotide arrays
Absolute mRNA concentrations from sequence-specific calibration of oligonucleotide arrays
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DOI：
10.1093/nar/gkg283
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发表时间：
2003-04-01
影响因子：
14.9
通讯作者：
Naef, F
中科院分区：
文献类型：
--
作者：
Hekstra, D;Taussig, AR;Naef, F
寡核苷酸微阵列基于标记的mRNA分子与玻璃表面上的短长度寡核苷酸探针的杂交。两种效应已被证明会影响原始数据：探针杂交特性的序列依赖性和表面吸附过程产生的化学饱和度。我们解决这两个问题，同时使用物理动机的杂交模型。公开的校准数据集的基础上，我们表明，朗缪尔吸附准确地描述基因芯片杂交，与模型参数，我们预测从探针的序列组成。由于这些参数具有物理单位，因此我们能够以皮摩尔估计绝对mRNA浓度。此外，通过考虑化学饱和度，我们大大减少了通常发生在高浓度的差异表达估计的压缩偏差。
Oligonucleotide microarrays are based on the hybridization of labeled mRNA molecules to short length oligonucleotide probes on a glass surface. Two effects have been shown to affect the raw data: the sequence dependence of the probe hybridization properties and the chemical saturation resulting from surface adsorption processes. We address both issues simultaneously using a physically motivated hybridization model. Based on publicly available calibration data sets, we show that Langmuir adsorption accurately describes GeneChip hybridization, with model parameters that we predict from the sequence composition of the probes. Because these parameters have physical units, we are able to estimate absolute mRNA concentrations in picomolar. Additionally, by accounting for chemical saturation, we substantially reduce the compressive bias of differential expression estimates that normally occurs toward high concentrations.