Exome RNA sequencing reveals rare and novel alternative transcripts.
Exome RNA sequencing reveals rare and novel alternative transcripts.
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DOI：
10.1093/nar/gks816
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发表时间：
2013-01-07
影响因子：
14.9
通讯作者：
Feuk L
中科院分区：
文献类型：
--
作者：
Halvardson J;Zaghlool A;Feuk L
RNA测序已经成为对全球基因表达进行无假设表征的重要方法。RNA测序的局限性之一是，大多数序列读取代表高表达的转录本，而低水平的转录本很难检测。为了将传统表达阵列的优点与RNA测序的优点结合起来，我们在总RNA测序之前使用了完整的外显子组浓缩。我们证明，整个外显子组捕获可以成功地应用于cDNA来研究人类组织中的转录图谱。通过引入外显子组浓缩步骤，我们能够识别存在于非常低水平的转录本，这低于传统RNA测序中的检测水平。尽管浓缩提高了检测转录本存在的能力，但它也降低了表达水平量化的准确性。我们的结果产生了大量新的外显子和剪接异构体，这表明传统的RNA测序方法只检测到完整转录多样性的一小部分。我们认为，RNA的整个外显子组丰富是一种适合于全基因组发现新的转录本、选择性剪接变异体和融合基因的策略。
RNA sequencing has become an important method to perform hypothesis-free characterization of global gene expression. One of the limitations of RNA sequencing is that most sequence reads represent highly expressed transcripts, whereas low level transcripts are challenging to detect. To combine the benefits of traditional expression arrays with the advantages of RNA sequencing, we have used whole exome enrichment prior to sequencing of total RNA. We show that whole exome capture can be successfully applied to cDNA to study the transcriptional landscape in human tissues. By introducing the exome enrichment step, we are able to identify transcripts present at very low levels, which are below the level of detection in conventional RNA sequencing. Although the enrichment increases the ability to detect presence of transcripts, it also lowers the accuracy of quantification of expression levels. Our results yield a large number of novel exons and splice isoforms, suggesting that conventional RNA sequencing methods only detect a small fraction of the full transcript diversity. We propose that whole exome enrichment of RNA is a suitable strategy for genome-wide discovery of novel transcripts, alternative splice variants and fusion genes.
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