Direct long-read RNA sequencing identifies a subset of questionable exitrons likely arising from reverse transcription artifacts.
Direct long-read RNA sequencing identifies a subset of questionable exitrons likely arising from reverse transcription artifacts.
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DOI：
10.1186/s13059-021-02411-1
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发表时间：
2021-06-28
期刊：
影响因子：
12.3
通讯作者：
Thomas-Tikhonenko A
中科院分区：
文献类型：
--
作者：
Schulz L;Torres-Diz M;Cortés-López M;Hayer KE;Asnani M;Tasian SK;Barash Y;Sotillo E;Zarnack K;König J;Thomas-Tikhonenko A
除其他因素外，淋巴细胞白血病对CD19定向免疫疗法的耐药性归因于几种异常的CD19前mrna剪接事件，包括最近报道的CD19外显子2内隐藏内含子的切除。虽然已知“外显子”存在于数百种人类转录本中，但我们发现，使用报告基因测定和直接长读RNA测序（RNA-seq）， CD19外显子是逆转录的产物。将我们的分析扩展到公开可用的数据集，我们发现了几十个可疑的外显子，被称为“假子”，它们只出现在dna -seq中，但从未出现在dna -seq中。我们的结果强调了dna -seq对转录物异构体验证的重要性。在线版本包含补充材料，可在10.1186/s13059-021-02411-1获得。
Resistance to CD19-directed immunotherapies in lymphoblastic leukemia has been attributed, among other factors, to several aberrant CD19 pre-mRNA splicing events, including recently reported excision of a cryptic intron embedded within CD19 exon 2. While “exitrons” are known to exist in hundreds of human transcripts, we discovered, using reporter assays and direct long-read RNA sequencing (dRNA-seq), that the CD19 exitron is an artifact of reverse transcription. Extending our analysis to publicly available datasets, we identified dozens of questionable exitrons, dubbed “falsitrons,” that appear only in cDNA-seq, but never in dRNA-seq. Our results highlight the importance of dRNA-seq for transcript isoform validation. The online version contains supplementary material available at 10.1186/s13059-021-02411-1.
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