Department of Organismic and Evolutionary Biology, Harvard University, Cambridge, Massachusetts, United States of America.
PLoS One. 2013 May 6;8(5):e61479. doi: 10.1371/journal.pone.0061479. Print 2013.
Most genomic resources available for insects represent the Holometabola, which are insects that undergo complete metamorphosis like beetles and flies. In contrast, the Hemimetabola (direct developing insects), representing the basal branches of the insect tree, have very few genomic resources. We have therefore created a large and publicly available transcriptome for the hemimetabolous insect Gryllus bimaculatus (cricket), a well-developed laboratory model organism whose potential for functional genetic experiments is currently limited by the absence of genomic resources. cDNA was prepared using mRNA obtained from adult ovaries containing all stages of oogenesis, and from embryo samples on each day of embryogenesis. Using 454 Titanium pyrosequencing, we sequenced over four million raw reads, and assembled them into 21,512 isotigs (predicted transcripts) and 120,805 singletons with an average coverage per base pair of 51.3. We annotated the transcriptome manually for over 400 conserved genes involved in embryonic patterning, gametogenesis, and signaling pathways. BLAST comparison of the transcriptome against the NCBI non-redundant protein database (nr) identified significant similarity to nr sequences for 55.5% of transcriptome sequences, and suggested that the transcriptome may contain 19,874 unique transcripts. For predicted transcripts without significant similarity to known sequences, we assessed their similarity to other orthopteran sequences, and determined that these transcripts contain recognizable protein domains, largely of unknown function. We created a searchable, web-based database to allow public access to all raw, assembled and annotated data. This database is to our knowledge the largest de novo assembled and annotated transcriptome resource available for any hemimetabolous insect. We therefore anticipate that these data will contribute significantly to more effective and higher-throughput deployment of molecular analysis tools in Gryllus.
大多数昆虫的基因组资源代表全变态昆虫，即经历完全变态的昆虫，如甲虫和苍蝇。相比之下，半变态昆虫（直接发育的昆虫）是昆虫树的基础分支，其基因组资源非常有限。因此，我们为半变态昆虫蟋蟀（Gryllus bimaculatus）创建了一个大型的、公开可用的转录组，蟋蟀是一种发育良好的实验室模式生物，其功能遗传实验的潜力目前受到基因组资源缺乏的限制。使用从包含卵发生各个阶段的成年卵巢和胚胎发生的每一天的胚胎样本中获得的 mRNA 制备 cDNA。使用 454 Titanium 焦磷酸测序，我们对超过四百万个原始读数进行了测序，并将它们组装成 21512 个同工基因（预测转录物）和 120805 个单基因，每个碱基对的平均覆盖率为 51.3。我们手动注释了转录组，涉及胚胎模式形成、配子发生和信号通路的 400 多个保守基因。将转录组与 NCBI 非冗余蛋白质数据库（nr）进行 BLAST 比较，确定了 55.5%的转录组序列与 nr 序列具有显著相似性，并表明转录组可能包含 19874 个独特的转录物。对于与已知序列没有显著相似性的预测转录物，我们评估了它们与其他直翅目序列的相似性，并确定这些转录物包含可识别的蛋白质结构域，主要功能未知。我们创建了一个可搜索的基于网络的数据库，允许公众访问所有原始、组装和注释的数据。据我们所知，这是任何半变态昆虫中最大的从头组装和注释转录组资源。因此，我们预计这些数据将极大地促进在蟋蟀中更有效地部署分子分析工具和更高通量的部署。
