Measure transcript integrity using RNA-seq data.
Measure transcript integrity using RNA-seq data.
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DOI：
10.1186/s12859-016-0922-z
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发表时间：
2016-02-03
影响因子：
3
通讯作者：
Kocher JP
中科院分区：
文献类型：
--
作者：
Wang L;Nie J;Sicotte H;Li Y;Eckel-Passow JE;Dasari S;Vedell PT;Barman P;Wang L;Weinshiboum R;Jen J;Huang H;Kohli M;Kocher JP
存储的具有病理信息和医疗记录的生物样本是转化医学研究的宝贵资源。然而，从存档的临床组织中提取的RNA通常被显著降解。RNA降解以基因特异性方式扭曲RNA-seq读段覆盖，并对全基因组基因表达谱产生深远影响。我们开发了转录完整性数（TIN）来测量RNA降解。当应用于3个独立的RNA-seq数据集时，我们证明了TIN是在转录物和样品水平上RNA降解的可靠和灵敏的量度。通过比较10个具有较低RNA完整性的前列腺癌临床样本与10个具有较高RNA质量的样本，我们证明了用TIN评分校准基因表达计数可以通过减少假阳性和恢复生物学上有意义的途径来有效地中和RNA降解效应。当使用测序质量控制联盟生成的RNA测序数据中的加标转录本进一步评估TIN校正的性能时，我们发现TIN校正可以更好地控制假阳性和假阴性（灵敏度= 0.89，特异性= 0.91，准确度= 0.90），与未进行TIN校正的基因表达分析结果相比（灵敏度= 0.98，特异度= 0.50，准确度= 0.86）。TIN是一种可靠的RNA完整性测量方法，可用于中和体外RNA降解效应和改善差异基因表达分析。本文的在线版本（doi：10.1186/s12859-016-0922-z）包含补充材料，可供授权用户使用。
Stored biological samples with pathology information and medical records are invaluable resources for translational medical research. However, RNAs extracted from the archived clinical tissues are often substantially degraded. RNA degradation distorts the RNA-seq read coverage in a gene-specific manner, and has profound influences on whole-genome gene expression profiling. We developed the transcript integrity number (TIN) to measure RNA degradation. When applied to 3 independent RNA-seq datasets, we demonstrated TIN is a reliable and sensitive measure of the RNA degradation at both transcript and sample level. Through comparing 10 prostate cancer clinical samples with lower RNA integrity to 10 samples with higher RNA quality, we demonstrated that calibrating gene expression counts with TIN scores could effectively neutralize RNA degradation effects by reducing false positives and recovering biologically meaningful pathways. When further evaluating the performance of TIN correction using spike-in transcripts in RNA-seq data generated from the Sequencing Quality Control consortium, we found TIN adjustment had better control of false positives and false negatives (sensitivity = 0.89, specificity = 0.91, accuracy = 0.90), as compared to gene expression analysis results without TIN correction (sensitivity = 0.98, specificity = 0.50, accuracy = 0.86). TIN is a reliable measurement of RNA integrity and a valuable approach used to neutralize in vitro RNA degradation effect and improve differential gene expression analysis. The online version of this article (doi:10.1186/s12859-016-0922-z) contains supplementary material, which is available to authorized users.