Characterization of Oncidium 'Gower Ramsey' Transcriptomes using 454 GS-FLX Pyrosequencing and Their Application to the Identification of Genes Associated with Flowering Time
Characterization of Oncidium 'Gower Ramsey' Transcriptomes using 454 GS-FLX Pyrosequencing and Their Application to the Identification of Genes Associated with Flowering Time
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DOI：
10.1093/pcp/pcr101
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发表时间：
2011-09-01
影响因子：
4.9
通讯作者：
Yang, Chang-Hsien
中科院分区：
文献类型：
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作者：
Chang, Yu-Yun;Chu, Yen-Wei;Yang, Chang-Hsien
文心兰是台湾花卉产业的一种有价值和成功的商业兰花。然而，目前还没有文心兰转录基因全序列的基因组参考，以促进分子生物学研究的发展和这些兰花的育种。在本研究中，我们构建了六个不同器官的文心兰cDNA文库：叶、假球茎、幼花、花序、花蕾和成熟花。我们利用454焦磷酸测序技术对文心兰转录组进行了高通量深度测序，获得了90万个读数，平均长度为328个碱基，总共有3.01亿个碱基。序列的从头组装从796,463个阅读数和120,219个单体中产生了50,908个重叠群序列，平均长度为493bp。用BLAST对组装的序列进行注释，并用TBLASTX分别与来自拟南芥和水稻基因组的12,757和13,931个Unigene转录本进行匹配。对带注释的文心兰重叠群进行的基因本体论(GO)分析表明，大多数测序的基因与未知的分子功能、细胞过程和细胞内成分有关。此外，该文库还获得了一个完整的开花相关表达序列，包括光周期途径中的大部分基因和15个具有保守CCT结构域的Constans-like(COL)同源物。这些数据表明，本研究中建立的文心兰表达序列标签(EST)数据库具有足够的覆盖率，可以作为研究兰花开花途径和其他各种生物学途径的工具。已经建立了一个文献兰基因组数据库网站，并可在网上公开使用(http://predictor.nchu.edu.tw/oogb/)
Oncidium 'Gower Ramsey' is a valuable and successful commercial orchid for the floriculture industry in Taiwan. However, no genome reference for entire sequences of the transcribed genes currently exists for Oncidium orchids, to facilitate the development of molecular biological studies and the breeding of these orchids. In this study, we generated Oncidium cDNA libraries for six different organs: leaves, pseudobulbs, young inflorescences, inflorescences, flower buds and mature flowers. We utilized 454-pyrosequencing technology to perform high-throughput deep sequencing of the Oncidium transcriptome, yielding > 0.9 million reads with an average length of 328 bp, for a total of 301 million bases. De novo assembly of the sequences yielded 50,908 contig sequences with an average length of 493 bp from 796,463 reads and 120,219 singletons. The assembled sequences were annotated using BLAST, and a total of 12,757 and 13,931 unigene transcripts from the Arabidopsis and rice genomes were matched by TBLASTX, respectively. A Gene Ontology (GO) analysis of the annotated Oncidium contigs revealed that the majority of sequenced genes were associated with 'unknown molecular function', 'cellular process' and 'intracellular components'. Furthermore, a complete flowering-associated expressed sequence that included most of the genes in the photoperiod pathway and the 15 CONSTANS-LIKE (COL) homologs with the conserved CCT domain was obtained in this collection. These data revealed that the Oncidium expressed sequence tag (EST) database generated in this study has sufficient coverage to be used as a tool to investigate the flowering pathway and various other biological pathways in orchids. An OncidiumOrchidGenomeBase (OOGB) website has been constructed and is publicly available online (http://predictor.nchu.edu.tw/oogb/)