Current issues for DNA microarrays: platform comparison, double linear amplification, and universal RNA reference
Current issues for DNA microarrays: platform comparison, double linear amplification, and universal RNA reference
复制标题
DOI：
10.1016/j.jbiotec.2004.05.006
复制
发表时间：
2004-09-09
影响因子：
4.1
通讯作者：
Choi, S
中科院分区：
文献类型：
--
作者：
Park, PJ;Cao, YA;Choi, S
DNA微阵列技术已被广泛用于同时测定数千个基因的表达水平。在该技术的实施和大量表达数据的分析中，已经使用了各种方法。然而，一些实际问题仍然没有得到解决，在一个令人满意的方式，其中最关键的是缺乏一致的结果，在不同的阵列平台。在这项研究中，我们提出了几个微阵列平台的比较[Affytek寡核苷酸阵列，自定义互补DNA（cDNA）阵列，和自定义寡核苷酸阵列打印的寡核苷酸从三个不同的来源]以及用于微阵列的目标制备和参考设计的各种方法的分析。结果表明，平台之间的表达水平的成对相关性总体上相对较低，但高表达基因的对数比是强相关的，特别是在AffyIteration和cDNA阵列之间。将微阵列测量结果与23个基因的定量实时聚合酶链反应（QRT-PCR）结果进行比较，并对每个平台的不同程度的一致性进行表征。我们还开发并测试了一种双重扩增方法，该方法允许使用较少量的起始材料。与与单轮扩增靶杂交的阵列相比，增加的扩增轮产生可再现的结果。终于来了对双通道微阵列使用通用RNA参考的可靠性进行了测试，结果表明使用相同对照的多个实验条件的比较是准确的。(C)2004 Elsevier B. V.保留所有权利。
DNA microarray technology has been widely used to simultaneously determine the expression levels of thousands of genes. A variety of approaches have been used, both in the implementation of this technology and in the analysis of the large amount of expression data. However, several practical issues still have not been resolved in a satisfactory manner, and among the most critical is the lack of agreement in the results obtained in different array platforms. In this study, we present a comparison of several microarray platforms [Affymetrix oligonucleotide arrays, custom complementary DNA (cDNA) arrays, and custom oligo arrays printed with oligonucleotides from three different sources] as well as analysis of various methods used for microarray target preparation and the reference design. The results indicate that the pairwise correlations of expression levels between platforms are relative low overall but that the log ratios of the highly expressed genes are strongly correlated, especially between Affymetrix and cDNA arrays. The microarray measurements were compared with quantitative real-time-polymerase chain reaction (QRT-PCR) results for 23 genes, and the varying degrees of agreement for each platform were characterized. We have also developed and tested a double amplification method which allows the use of smaller amounts of starting material. The added round of amplification produced reproducible results as compared to the arrays hybridized with single round amplified targets. Finally. the reliability of using a universal RNA reference for two-channel microarrays was tested and the results suggest that comparisons of multiple experimental conditions using the same control can be accurate. (C) 2004 Elsevier B.V. All rights reserved.