Deep Seq, Faculty of Medicine and Health Sciences, Queen's Medical Centre, University of Nottingham, Nottingham, United Kingdom.
PLoS One. 2012;7(3):e33784. doi: 10.1371/journal.pone.0033784. Epub 2012 Mar 20.
Understanding the genetic and evolutionary basis of animal morphological diversity will require comparative developmental studies that use new model organisms. This necessitates development of tools for the study of genetics and also the generation of sequence information of the organism to be studied. The development of next generation sequencing technology has enabled quick and cost effective generation of sequence information. Parhyale hawaiensis has emerged as a model organism of choice due to the development of advanced molecular tools, thus P. hawaiensis genetic information will help drive functional studies in this organism.Here we present a transcriptome and miRNA collection generated using next generation sequencing platforms. We generated approximately 1.7 million reads from a P. hawaiensis cDNA library constructed from embryos up to the germ band stage. These reads were assembled into a dataset comprising 163,501 transcripts.Using the combined annotation of Annot8r and pfam2go, Gene Ontology classifications was assigned to 20,597 transcripts. Annot8r was used to provide KEGG orthology to our transcript dataset. A total of 25,292 KEGG pathway assignments were defined and further confirmed with reciprocal blast against the NCBI nr protein database. This has identified many P. hawaiensis gene orthologs of key conserved signalling pathways involved in development. We also generated small RNA sequences from P. hawaiensis, identifying 55 conserved miRNAs. Sequenced small RNAs that were not annotated by stringent comparison to mirBase were used to search the Daphnia pulex for possible novel miRNAs. Using a conservative approach, we have identified 51 possible miRNA candidates conserved in the Daphnia pulex genome, which could be potential crustacean/arthropod specific miRNAs. Our study presents gene and miRNA discovery in a new model organism that does not have a sequenced genome. The data provided by our work will be valuable for the P. hawaiensis community as well as the wider evolutionary developmental biology community.
理解动物形态多样性的遗传和进化基础将需要使用新的模式生物进行比较发育研究。这需要开发用于研究遗传学的工具，以及生成要研究的生物体的序列信息。下一代测序技术的发展使得快速且具有成本效益的序列信息生成成为可能。由于先进的分子工具的发展，Parhyale hawaiensis 已成为首选的模式生物，因此 P. hawaiensis 的遗传信息将有助于推动该生物体的功能研究。在这里，我们展示了使用下一代测序平台生成的转录组和 miRNA 集合。我们从胚胎到原肠胚阶段构建的 P. hawaiensis cDNA 文库中生成了大约 170 万个读数。这些读数被组装成一个包含 163501 个转录本的数据集。使用 Annot8r 和 pfam2go 的联合注释，将基因本体论分类分配给 20597 个转录本。Annot8r 用于为我们的转录本数据集提供 KEGG 直系同源物。总共定义了 25292 个 KEGG 途径分配，并通过与 NCBI nr 蛋白质数据库的反向 Blast 进一步确认。这确定了许多参与发育的关键保守信号通路的 P. hawaiensis 基因直系同源物。我们还从 P. hawaiensis 中生成了小 RNA 序列，鉴定出 55 个保守的 miRNA。通过与 mirBase 进行严格比较未注释的测序小 RNA 被用于在 Daphnia pulex 中搜索可能的新 miRNA。使用保守方法，我们在 Daphnia pulex 基因组中鉴定了 51 个可能保守的 miRNA 候选物，它们可能是潜在的甲壳类/节肢动物特异性 miRNA。我们的研究在没有测序基因组的新模型生物中进行了基因和 miRNA 的发现。我们工作提供的数据对于 P. hawaiensis 社区以及更广泛的进化发育生物学社区都将具有重要价值。
