Systematic analysis of transcribed loci in ENCODE regions using RACE sequencing reveals extensive transcription in the human genome
Systematic analysis of transcribed loci in ENCODE regions using RACE sequencing reveals extensive transcription in the human genome
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DOI：
10.1186/gb-2008-9-1-r3
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发表时间：
2008-01-01
期刊：
影响因子：
12.3
通讯作者：
Snyder, Michael
中科院分区：
文献类型：
--
作者：
QianWu, Jia;Du, Jiang;Snyder, Michael
背景：最近对哺乳动物转录组的研究揭示了转录多样性中的大量额外转录区域和异常复杂的转录多样性。然而，关于基因组的哪个部分被转录，这些新转录本的确切结构，以及转录本的水平仍然存在很大的不确定性。结果：我们使用快速扩增基因末端扩增(RACE)测序的方法查询了420个选定的DNA元素百科全书(Encode)区域的转录位点。我们分析了已注释的已知基因区域，但主要集中在新的转录活性区域(TAR)，这些区域以前是通过高密度寡核苷酸拼接阵列识别的，以及不被认为是转录的随机区域。我们发现RACE测序非常敏感，能够检测到微阵列无法检测到的特定细胞类型中的低水平转录本。我们还观察到了许多正义-反义转录本的例子；进一步的分析表明，许多反义转录本(但不是全部)可能是逆转录反应产生的产物。我们的结果表明，所分析的大多数新的TAR(60%)与其他新的TAR或已知的外显子有关。在之前没有注释的随机区域中，17%的区域显示出重叠的转录本。此外，据估计，9%的新转录本编码蛋白质。结论：RACE测序是一种有效、敏感和高精度的方法来表征特定细胞/组织类型的转录组。使用这种方法，似乎有很大一部分基因组是以PolA+RNA表示的。此外，新RNA的一小部分可以编码蛋白质，并可能具有功能。
Background: Recent studies of the mammalian transcriptome have revealed a large number of additional transcribed regions and extraordinary complexity in transcript diversity. However, there is still much uncertainty regarding precisely what portion of the genome is transcribed, the exact structures of these novel transcripts, and the levels of the transcripts produced.Results: We have interrogated the transcribed loci in 420 selected ENCyclopedia Of DNA Elements (ENCODE) regions using rapid amplification of cDNA ends (RACE) sequencing. We analyzed annotated known gene regions, but primarily we focused on novel transcriptionally active regions (TARs), which were previously identified by high-density oligonucleotide tiling arrays and on random regions that were not believed to be transcribed. We found RACE sequencing to be very sensitive and were able to detect low levels of transcripts in specific cell types that were not detectable by microarrays. We also observed many instances of sense-antisense transcripts; further analysis suggests that many of the antisense transcripts (but not all) may be artifacts generated from the reverse transcription reaction. Our results show that the majority of the novel TARs analyzed (60%) are connected to other novel TARs or known exons. Of previously unannotated random regions, 17% were shown to produce overlapping transcripts. Furthermore, it is estimated that 9% of the novel transcripts encode proteins.Conclusion: We conclude that RACE sequencing is an efficient, sensitive, and highly accurate method for characterization of the transcriptome of specific cell/tissue types. Using this method, it appears that much of the genome is represented in polyA+ RNA. Moreover, a fraction of the novel RNAs can encode protein and are likely to be functional.