RNA-Seq of Arabidopsis Pollen Uncovers Novel Transcription and Alternative Splicing
RNA-Seq of Arabidopsis Pollen Uncovers Novel Transcription and Alternative Splicing
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DOI：
10.1104/pp.112.211441
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发表时间：
2013-06-01
期刊：
影响因子：
7.4
通讯作者：
Qin, Peng
中科院分区：
文献类型：
--
作者：
Loraine, Ann E.;McCormick, Sheila;Qin, Peng
拟南芥的花粉粒含有两个被包围在一个单倍体营养细胞中的单倍体精细胞。萌发时，营养细胞挤出花粉管，花粉管携带精子到胚珠受精。了解花粉转录本的身份、相对丰度和剪接模式将有助于我们更好地了解花粉，并有助于研究植物中的组织特异性剪接。大多数拟南芥花粉转录组研究使用ATH 1微阵列，它不分析剪接变异体，缺乏许多基因的特异性探针组。为了研究花粉转录组，我们进行了拟南芥花粉和幼苗的高通量测序（RNA-Seq）进行比较。基因表达在幼苗中更多样化，而参与细胞壁生物发生的基因在花粉中高表达。RNA-Seq检测到花粉中表达的至少4，172个蛋白质编码基因，包括仅通过非特异性探针组测定的289个。额外的外显子和以前未注释的59和39花粉表达基因的非翻译区被发现。我们在基因组中检测到以前没有注释为表达的区域; 14个进行了测试，12个通过聚合酶链反应进行了确认。空位读段比对揭示了由10个或更多个剪接读段比对支持的1，908个高置信度新剪接事件。花粉和幼苗中的选择性剪接模式高度相关。对于大多数选择性剪接的基因，除了一些参与RNA剪接的编码蛋白外，花粉和幼苗中的变体比例相似。这项研究强调了植物中剪接模式的鲁棒性，以及使用集成基因组浏览器等交互式工具对RNA-Seq数据进行注释和可视化的重要性。
Pollen grains of Arabidopsis (Arabidopsis thaliana) contain two haploid sperm cells enclosed in a haploid vegetative cell. Upon germination, the vegetative cell extrudes a pollen tube that carries the sperm to an ovule for fertilization. Knowing the identity, relative abundance, and splicing patterns of pollen transcripts will improve our understanding of pollen and allow investigation of tissue-specific splicing in plants. Most Arabidopsis pollen transcriptome studies have used the ATH1 microarray, which does not assay splice variants and lacks specific probe sets for many genes. To investigate the pollen transcriptome, we performed high-throughput sequencing (RNA-Seq) of Arabidopsis pollen and seedlings for comparison. Gene expression was more diverse in seedling, and genes involved in cell wall biogenesis were highly expressed in pollen. RNA-Seq detected at least 4,172 protein-coding genes expressed in pollen, including 289 assayed only by nonspecific probe sets. Additional exons and previously unannotated 59 and 39 untranslated regions for pollen-expressed genes were revealed. We detected regions in the genome not previously annotated as expressed; 14 were tested and 12 were confirmed by polymerase chain reaction. Gapped read alignments revealed 1,908 high-confidence new splicing events supported by 10 or more spliced read alignments. Alternative splicing patterns in pollen and seedling were highly correlated. For most alternatively spliced genes, the ratio of variants in pollen and seedling was similar, except for some encoding proteins involved in RNA splicing. This study highlights the robustness of splicing patterns in plants and the importance of ongoing annotation and visualization of RNA-Seq data using interactive tools such as Integrated Genome Browser.