BaRTv1.0: an improved barley reference transcript dataset to determine accurate changes in the barley transcriptome using RNA-seq
BaRTv1.0: an improved barley reference transcript dataset to determine accurate changes in the barley transcriptome using RNA-seq
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DOI：
10.1186/s12864-019-6243-7
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发表时间：
2019-12-11
期刊：
影响因子：
4.4
通讯作者：
Simpson, Craig G.
中科院分区：
文献类型：
--
作者：
Rapazote-Flores, Paulo;Bayer, Micha;Simpson, Craig G.
背景资料：由于计算组装、基因表达定量和剪接分析的离散步骤，分析RNA-seq数据所需的时间差异很大。最近的快速非比对工具，如Kallisto和鲑鱼克服了这些问题，但这些工具需要一个高质量的，全面的参考转录本数据集（RTD），这是很少在plants.Results：一个高质量的，非冗余的大麦基因RTD和数据库（大麦参考转录本- BaRTv1.0）已被生成。BaRTv1.0，构建了一系列的组织，品种和非生物处理和转录组装和对齐的大麦品种。Morex参考基因组（Mascher等人Nature; 544：427-433，2017）。来自大麦品种Haruna nijo的全长cDNA（松本（Matsumoto）等人，植物生理学（Plant Physiol）; 156：20-28，2011）确定了转录物覆盖率，并且高分辨率RT-PCR验证了五种不同器官和组织中86个基因的选择性剪接（AS）转录物。这些方法被用作基准选择最佳的大麦RTD。还进行了BaRTv1.0-选择性剪接异构体的定量（QUASI）以克服由于转录物的5'和3' UTR末端的变化而导致的不准确定量。BaRTv1.0-QUASI用于五个大麦器官/组织的RNA-seq数据的准确转录本定量。该分析确定了20，972个显著差异表达的基因，2791个差异可变剪接基因和2768个具有差异转录本use.Conclusion转录本数据集由60，444个基因和177，240个转录本组成。与目前的大麦转录本相比，BaRTv1.0转录本通常更长，具有更少的片段化和改进的基因模型，这些模型得到剪接点读数的很好支持。使用BaRTv1.0进行精确的转录本定量可以对基因表达和AS进行常规分析。
Background: The time required to analyse RNA-seq data varies considerably, due to discrete steps for computational assembly, quantification of gene expression and splicing analysis. Recent fast non-alignment tools such as Kallisto and Salmon overcome these problems, but these tools require a high quality, comprehensive reference transcripts dataset (RTD), which are rarely available in plants.Results: A high-quality, non-redundant barley gene RTD and database (Barley Reference Transcripts - BaRTv1.0) has been generated. BaRTv1.0, was constructed from a range of tissues, cultivars and abiotic treatments and transcripts assembled and aligned to the barley cv. Morex reference genome (Mascher et al. Nature; 544: 427-433, 2017). Full-length cDNAs from the barley variety Haruna nijo (Matsumoto et al. Plant Physiol; 156: 20-28, 2011) determined transcript coverage, and high-resolution RT-PCR validated alternatively spliced (AS) transcripts of 86 genes in five different organs and tissue. These methods were used as benchmarks to select an optimal barley RTD. BaRTv1.0-Quantification of Alternatively Spliced Isoforms (QUASI) was also made to overcome inaccurate quantification due to variation in 5' and 3' UTR ends of transcripts. BaRTv1.0-QUASI was used for accurate transcript quantification of RNA-seq data of five barley organs/tissues. This analysis identified 20,972 significant differentially expressed genes, 2791 differentially alternatively spliced genes and 2768 transcripts with differential transcript usage.Conclusion: A high confidence barley reference transcript dataset consisting of 60,444 genes with 177,240 transcripts has been generated. Compared to current barley transcripts, BaRTv1.0 transcripts are generally longer, have less fragmentation and improved gene models that are well supported by splice junction reads. Precise transcript quantification using BaRTv1.0 allows routine analysis of gene expression and AS.