TERA-Seq: true end-to-end sequencing of native RNA molecules for transcriptome characterization.
TERA-Seq: true end-to-end sequencing of native RNA molecules for transcriptome characterization.
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DOI：
10.1093/nar/gkab713
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发表时间：
2021-11-18
影响因子：
14.9
通讯作者：
Mourelatos Z
中科院分区：
文献类型：
--
作者：
Ibrahim F;Oppelt J;Maragkakis M;Mourelatos Z
通过纳米孔对单个天然RNA分子进行直接测序具有很强的潜力，可以改变RNA生物学和临床诊断各个方面的研究。Oxford Nanopore Technologies的现有平台无法对RNA的5′端进行测序，并且仅限于聚腺苷酸化分子。在这里，我们开发了真正的端到端RNA测序（TERA-Seq），这是一个解决这些限制的平台，可以进行更彻底的转录组表征。TERA-Seq描述了多聚腺苷酸化和非多聚腺苷酸化的RNA分子，并通过连接独特设计的衔接子（与转录本一起沿着测序）准确识别其天然5 '和3'末端。我们发现人类细胞中的加帽全长mRNA在单分子水平上显示出poly（A）尾长度的显著变化。我们报告普遍加盖下游的典型转录起始位点，否则完全剪接和多聚腺苷酸化分子。我们在单分子水平上揭示了RNA的加工和衰变，发现mRNA的衰变通常是从5′端开始的，同时经常保留poly（A）尾。TERA-Seq将证明在许多应用中是有用的，其中需要单个天然RNA分子及其同种型的真正端到端直接测序。
Direct sequencing of single, native RNA molecules through nanopores has a strong potential to transform research in all aspects of RNA biology and clinical diagnostics. The existing platform from Oxford Nanopore Technologies is unable to sequence the very 5′ ends of RNAs and is limited to polyadenylated molecules. Here, we develop True End-to-end RNA Sequencing (TERA-Seq), a platform that addresses these limitations, permitting more thorough transcriptome characterization. TERA-Seq describes both poly- and non-polyadenylated RNA molecules and accurately identifies their native 5′ and 3′ ends by ligating uniquely designed adapters that are sequenced along with the transcript. We find that capped, full-length mRNAs in human cells show marked variation of poly(A) tail lengths at the single molecule level. We report prevalent capping downstream of canonical transcriptional start sites in otherwise fully spliced and polyadenylated molecules. We reveal RNA processing and decay at single molecule level and find that mRNAs decay cotranslationally, often from their 5′ ends, while frequently retaining poly(A) tails. TERA-Seq will prove useful in many applications where true end-to-end direct sequencing of single, native RNA molecules and their isoforms is desirable.
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影响因子：
48
作者：
Huber W;Carey VJ;Gentleman R;Anders S;Carlson M;Carvalho BS;Bravo HC;Davis S;Gatto L;Girke T;Gottardo R;Hahne F;Hansen KD;Irizarry RA;Lawrence M;Love MI;MacDonald J;Obenchain V;Oleś AK;Pagès H;Reyes A;Shannon P;Smyth GK;Tenenbaum D;Waldron L;Morgan M
通讯作者：
Morgan M
影响因子：
64.8
作者：
通讯作者：
--
影响因子：
9.2
作者：
Addo-Quaye, Charles;Eshoo, Tifani W.;Axtell, Michael J.
通讯作者：
Axtell, Michael J.
影响因子：
11.8
作者：
Gregory, Brian D.;O'Malley, Ronan C.;Ecker, Joseph R.
通讯作者：
Ecker, Joseph R.
影响因子：
14.8
作者：
Ibrahim, Fadia;Mourelatos, Zissimos
通讯作者：
Mourelatos, Zissimos