Tests of rRNA hybridization to microarrays suggest that hybridization characteristics of oligonucleotide probes for species discrimination cannot be predicted.
Tests of rRNA hybridization to microarrays suggest that hybridization characteristics of oligonucleotide probes for species discrimination cannot be predicted.
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RRNA与微阵列的杂交测试表明，无法预测寡核苷酸探针对物种歧视的杂交特征。
DOI：
10.1093/nar/gkl133
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发表时间：
2006-05-17
影响因子：
14.9
通讯作者：
Tautz D
中科院分区：
文献类型：
--
作者：
Pozhitkov A;Noble PA;Domazet-Loso T;Nolte AW;Sonnenberg R;Staehler P;Beier M;Tautz D
rRNA与基因芯片杂交是一种很有前途的原核和真核生物物种鉴定方法。通常，通过荧光强度测量结合靶的量，并且假定信号强度与靶浓度直接相关。使用13种不同的真核LSU rRNA靶序列和7693个短的完美匹配寡核苷酸探针，我们通过比较吉布斯自由能（ΔG°）计算结果与实验结果，评估了当前预测信号强度的方法。我们的评估显示，预测和实际强度之间的统计关系较差。尽管给定靶标的信号强度变化高达70倍，但没有一个预测因子能够完全解释这种变化。此外，没有组合不同的自由能条款，评估主成分和神经网络分析，提供了一个可靠的预测杂交效率。我们还研究了单碱基对错配（MM）（所有可能的类型和位置）对双链体信号强度的影响。我们发现，MM效应不同于从基于溶液的杂交预测的那些。这些结果建议对探针设计软件工具，使用热力学参数来评估探针质量的物种鉴定的应用程序。我们的研究结果意味着，寡核苷酸杂交的热力学性质是迄今为止还不清楚。
Hybridization of rRNAs to microarrays is a promising approach for prokaryotic and eukaryotic species identification. Typically, the amount of bound target is measured by fluorescent intensity and it is assumed that the signal intensity is directly related to the target concentration. Using thirteen different eukaryotic LSU rRNA target sequences and 7693 short perfect match oligonucleotide probes, we have assessed current approaches for predicting signal intensities by comparing Gibbs free energy (ΔG°) calculations to experimental results. Our evaluation revealed a poor statistical relationship between predicted and actual intensities. Although signal intensities for a given target varied up to 70-fold, none of the predictors were able to fully explain this variation. Also, no combination of different free energy terms, as assessed by principal component and neural network analyses, provided a reliable predictor of hybridization efficiency. We also examined the effects of single-base pair mismatch (MM) (all possible types and positions) on signal intensities of duplexes. We found that the MM effects differ from those that were predicted from solution-based hybridizations. These results recommend against the application of probe design software tools that use thermodynamic parameters to assess probe quality for species identification. Our results imply that the thermodynamic properties of oligonucleotide hybridization are by far not yet understood.
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DOI：
10.1073/pnas.1534744100
发表时间：
2003-09-30
影响因子：
11.1
作者：
Mei, R;Hubbell, E;Webster, TA
通讯作者：
Webster, TA
影响因子：
14.9
作者：
Peterson, AW;Heaton, RJ;Georgiadis, RM
通讯作者：
Georgiadis, RM
影响因子：
2.9
作者：
MARKY, LA;BRESLAUER, KJ
通讯作者：
BRESLAUER, KJ
影响因子：
14.9
作者：
Matveeva, OV;Shabalina, SA;Atkins, JF
通讯作者：
Atkins, JF
影响因子：
4.4
作者：
Noble, PA;Almeida, JS;Lovell, CR
通讯作者：
Lovell, CR