A high-resolution single-molecule sequencing-based Arabidopsis transcriptome using novel methods of Iso-seq analysis.
A high-resolution single-molecule sequencing-based Arabidopsis transcriptome using novel methods of Iso-seq analysis.
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DOI：
10.1186/s13059-022-02711-0
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发表时间：
2022-07-07
期刊：
影响因子：
12.3
通讯作者：
中科院分区：
文献类型：
--
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准确、全面的转录本序列注释是进行转录本定量、差异基因和转录本表达分析的基础。单分子长读测序技术提高了转录结构的完整性，包括选择性剪接、转录起始和多聚腺苷化位点。然而，测序错误、信使核糖核酸降解或不完整的cdna合成显著影响了精确度。我们提出了一个新的和全面的拟南芥参考转录数据集3(AtRTD3)。AtRTD3包含超过169,000个转录本--是目前最好的拟南芥转录组的两倍，其中包括1500多个新基因。78%的转录本来自具有准确定义的剪接连接和转录起始和结束位点的Iso-seq。我们开发了新的方法来准确地确定剪接连接和转录起始和结束位点。接头连接周围的不匹配纵断面提供了一项强大的功能，可以区分正确的接头连接并删除错误的接头连接。分层方法识别高置信度转录起始和结束位置，并删除由于降解而产生的片断转录。对拟南芥冷反应RNA-seq时间序列的分析表明，AtRTD3是对现有转录本的重大改进。AtRTD3为转录表达谱提供了更高的分辨率，并识别了冷诱导的差异转录开始和多聚腺苷化位点的使用。AtRTD3是目前拟南芥中功能最全面的转录组。它提高了从RNA-SEQ数据进行差异基因和转录本表达、差异选择性剪接和转录起始/结束位点使用分析的精确度。识别准确剪接连接和转录起始/结束位点的新方法具有广泛的适用性，并将改进任何物种的单分子测序分析。网上版载有补充材料，可在10.1186/s13059-022-02711-0查阅。
Accurate and comprehensive annotation of transcript sequences is essential for transcript quantification and differential gene and transcript expression analysis. Single-molecule long-read sequencing technologies provide improved integrity of transcript structures including alternative splicing, and transcription start and polyadenylation sites. However, accuracy is significantly affected by sequencing errors, mRNA degradation, or incomplete cDNA synthesis. We present a new and comprehensive Arabidopsis thaliana Reference Transcript Dataset 3 (AtRTD3). AtRTD3 contains over 169,000 transcripts—twice that of the best current Arabidopsis transcriptome and including over 1500 novel genes. Seventy-eight percent of transcripts are from Iso-seq with accurately defined splice junctions and transcription start and end sites. We develop novel methods to determine splice junctions and transcription start and end sites accurately. Mismatch profiles around splice junctions provide a powerful feature to distinguish correct splice junctions and remove false splice junctions. Stratified approaches identify high-confidence transcription start and end sites and remove fragmentary transcripts due to degradation. AtRTD3 is a major improvement over existing transcriptomes as demonstrated by analysis of an Arabidopsis cold response RNA-seq time-series. AtRTD3 provides higher resolution of transcript expression profiling and identifies cold-induced differential transcription start and polyadenylation site usage. AtRTD3 is the most comprehensive Arabidopsis transcriptome currently. It improves the precision of differential gene and transcript expression, differential alternative splicing, and transcription start/end site usage analysis from RNA-seq data. The novel methods for identifying accurate splice junctions and transcription start/end sites are widely applicable and will improve single-molecule sequencing analysis from any species. The online version contains supplementary material available at 10.1186/s13059-022-02711-0.
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DOI：
10.1093/bioinformatics/btx411
发表时间：
2017-10-15
期刊：
Bioinformatics (Oxford, England)
影响因子：
--
作者：
Guo W;Calixto CPG;Brown JWS;Zhang R
通讯作者：
Zhang R
影响因子：
16.6
作者：
Abdel-Ghany SE;Hamilton M;Jacobi JL;Ngam P;Devitt N;Schilkey F;Ben-Hur A;Reddy AS
通讯作者：
Reddy AS
通过单分子长读长测序分析红三叶草（Trifolium pratense L.）的转录本和剪接亚型
影响因子：
5.3
作者：
Chao Y;Yuan J;Li S;Jia S;Han L;Xu L
通讯作者：
Xu L
影响因子：
11.8
作者：
Hornyik, Csaba;Terzi, Lionel C.;Simpson, Gordon G.
通讯作者：
Simpson, Gordon G.
影响因子：
7
作者：
Braunschweig U;Barbosa-Morais NL;Pan Q;Nachman EN;Alipanahi B;Gonatopoulos-Pournatzis T;Frey B;Irimia M;Blencowe BJ
通讯作者：
Blencowe BJ