Romberg Tiburon Center and Department of Biology, San Francisco State University, Tiburon, California, United States of America.
PLoS One. 2010 Feb 19;5(2):e9327. doi: 10.1371/journal.pone.0009327.
BACKGROUND: With the emergence of a completed genome sequence of the freshwater crustacean Daphnia pulex, construction of genomic-scale sequence databases for additional crustacean sequences are important for comparative genomics and annotation. Porcelain crabs, genus Petrolisthes, have been powerful crustacean models for environmental and evolutionary physiology with respect to thermal adaptation and understanding responses of marine organisms to climate change. Here, we present a large-scale EST sequencing and cDNA microarray database project for the porcelain crab Petrolisthes cinctipes. METHODOLOGY/PRINCIPAL FINDINGS: A set of approximately 30K unique sequences (UniSeqs) representing approximately 19K clusters were generated from approximately 98K high quality ESTs from a set of tissue specific non-normalized and mixed-tissue normalized cDNA libraries from the porcelain crab Petrolisthes cinctipes. Homology for each UniSeq was assessed using BLAST, InterProScan, GO and KEGG database searches. Approximately 66% of the UniSeqs had homology in at least one of the databases. All EST and UniSeq sequences along with annotation results and coordinated cDNA microarray datasets have been made publicly accessible at the Porcelain Crab Array Database (PCAD), a feature-enriched version of the Stanford and Longhorn Array Databases. CONCLUSIONS/SIGNIFICANCE: The EST project presented here represents the third largest sequencing effort for any crustacean, and the largest effort for any crab species. Our assembly and clustering results suggest that our porcelain crab EST data set is equally diverse to the much larger EST set generated in the Daphnia pulex genome sequencing project, and thus will be an important resource to the Daphnia research community. Our homology results support the pancrustacea hypothesis and suggest that Malacostraca may be ancestral to Branchiopoda and Hexapoda. Our results also suggest that our cDNA microarrays cover as much of the transcriptome as can reasonably be captured in EST library sequencing approaches, and thus represent a rich resource for studies of environmental genomics.
背景:随着淡水甲壳动物溞属基因组序列的完成,构建其他甲壳动物基因组规模的序列数据库对于比较基因组学和注释非常重要。瓷蟹属的瓷蟹在环境和进化生理学方面,特别是在热适应和理解海洋生物对气候变化的反应方面,一直是强有力的甲壳动物模型。在这里,我们提出了一个大规模的 EST 测序和 cDNA 微阵列数据库项目,用于研究瓷蟹 Petrolisthes cinctipes。
方法/主要发现:从瓷蟹 Petrolisthes cinctipes 的一组组织特异性非归一化和混合组织归一化 cDNA 文库中,生成了一组约 98,000 个高质量 EST 序列,代表了大约 30,000 个独特序列(UniSeqs),这些序列大约代表了 19,000 个聚类。使用 BLAST、InterProScan、GO 和 KEGG 数据库搜索来评估每个 UniSeq 的同源性。大约 66%的 UniSeqs 在至少一个数据库中具有同源性。所有的 EST 和 UniSeq 序列以及注释结果和协调的 cDNA 微阵列数据集都可以在瓷蟹基因表达谱数据库(PCAD)中公开访问,这是斯坦福和长角牛基因表达谱数据库的一个功能丰富的版本。
结论/意义:这里呈现的 EST 项目代表了任何甲壳动物中第三大测序工作,也是任何蟹类中最大的测序工作。我们的组装和聚类结果表明,我们的瓷蟹 EST 数据集与在溞属基因组测序项目中生成的更大的 EST 数据集同样多样化,因此将成为溞属研究社区的一个重要资源。我们的同源性结果支持泛甲壳动物假说,并表明软甲纲可能是鳃足纲和六足纲的祖先。我们的结果还表明,我们的 cDNA 微阵列涵盖了转录组的大部分,这在 EST 文库测序方法中是可以合理捕获的,因此代表了环境基因组学研究的丰富资源。
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