Clark Tyson A, Schweitzer Anthony C, Chen Tina X, Staples Michelle K, Lu Gang, Wang Hui, Williams Alan, Blume John E
Affymetrix, Inc, Santa Clara, CA 95051, USA.
Genome Biol. 2007;8(4):R64. doi: 10.1186/gb-2007-8-4-r64.
Higher eukaryotes express a diverse population of messenger RNAs generated by alternative splicing. Large-scale methods for monitoring gene expression must adapt in order to accurately detect the transcript variation generated by this splicing.
We have designed a high-density oligonucleotide microarray with probesets for more than one million annotated and predicted exons in the human genome. Using these arrays and a simple algorithm that normalizes exon signal to signal from the gene as a whole, we have identified tissue-specific exons from a panel of 16 different normal adult tissues. RT-PCR validation confirms approximately 86% of the predicted tissue-enriched probesets. Pair-wise comparisons between the tissues suggest that as many as 73% of detected genes are differentially alternatively spliced. We also demonstrate how an inclusive exon microarray can be used to discover novel alternative splicing events. As examples, 17 new tissue-specific exons from 11 genes were validated by RT-PCR and sequencing.
In conjunction with a conceptually simple algorithm, comprehensive exon microarrays can detect tissue-specific alternative splicing events. Our data suggest significant expression outside of known exons and well annotated genes and a high frequency of alternative splicing events. In addition, we identified and validated a number of novel exons with tissue-specific splicing patterns. The tissue map data will likely serve as a valuable source of information on the regulation of alternative splicing.
高等真核生物表达由可变剪接产生的多种信使核糖核酸群体。用于监测基因表达的大规模方法必须做出调整，以便准确检测由这种剪接产生的转录本变异。
我们设计了一种高密度寡核苷酸微阵列，其探针集针对人类基因组中超过一百万个注释和预测的外显子。使用这些阵列和一种将外显子信号与整个基因的信号进行归一化的简单算法，我们从一组16种不同的正常成人组织中鉴定出了组织特异性外显子。逆转录聚合酶链反应验证确认了大约86%的预测组织富集探针集。组织之间的成对比较表明，多达73%的检测基因存在差异可变剪接。我们还展示了如何使用包含性外显子微阵列来发现新的可变剪接事件。例如，通过逆转录聚合酶链反应和测序验证了来自11个基因的17个新的组织特异性外显子。
结合一种概念上简单的算法，全面的外显子微阵列可以检测组织特异性可变剪接事件。我们的数据表明在已知外显子和注释良好的基因之外存在显著表达，并且可变剪接事件的频率很高。此外，我们鉴定并验证了许多具有组织特异性剪接模式的新外显子。组织图谱数据可能会成为可变剪接调控信息的宝贵来源。
