Data from: Transcriptome resources for the frogs Lithobates clamitans and Pseudacris regilla, emphasizing antimicrobial peptides and conserved loci for phylogenetics

We developed genetic resources for two North American frogs, Lithobates clamitans and Pseudacris regilla, widespread native amphibians that are potential indicator species of environmental health. For both species, mRNA from multiple tissues was sequenced using 454 technology. De novo assemblies with Mira3 resulted in 50,238 contigs (N50 = 687 bp) and 48,213 contigs (N50 = 686 bp) for L. clamitans and P. regilla, respectively, after clustering with CD-Hit-EST and purging contigs below 200 bp. We performed BLASTX similarity searches against the Xenopus tropicalis proteome and, for predicted ORFs, HMMER similarity searches against the Pfam-A database. Because there is broad interest in amphibian immune factors, we manually annotated putative antimicrobial peptides. To identify conserved regions suitable for amplicon re-sequencing across a broad taxonomic range, we performed an additional assembly of public short-read transcriptome data derived from two species of the genus Rana and identified reciprocal best TBLASTX matches among all assemblies. Although P. regilla, a hylid frog, is substantially more diverged from the ranid species, we identified 56 genes that were sufficiently conserved to allow non-degenerate primer design with Primer3. In addition to providing a foundation for comparative genomics and quantitative gene expression analysis, our results enable quick development of nuclear sequence-based markers for phylogenetics or population genetics.

本研究针对两种北美本土两栖类——青铜蛙（Lithobates clamitans）与太平洋树蛙（Pseudacris regilla）开发了遗传资源，二者均为广泛分布的本土两栖动物，是环境健康的潜在指示物种。针对这两个物种，我们采用454测序技术对多组织的mRNA进行了测序。使用Mira3进行从头组装后，通过CD-Hit-EST完成聚类并过滤掉长度低于200 bp的重叠群（contig），最终青铜蛙得到50238条重叠群，其N50值（N50）为687 bp；太平洋树蛙得到48213条重叠群，N50值（N50）为686 bp。我们针对热带爪蟾（Xenopus tropicalis）蛋白质组开展了BLASTX相似性搜索，并针对预测得到的开放阅读框（Open Reading Frame, ORF），利用HMMER软件对Pfam-A数据库进行了相似性检索。鉴于两栖动物免疫因子广受关注，我们对推定的抗菌肽进行了手动注释。为了鉴定适用于宽泛分类群的扩增子重测序保守区域，我们额外组装了来自蛙属（Rana）两个物种的公共短读长转录组数据，并在所有组装结果间鉴定出双向最优TBLASTX匹配。尽管太平洋树蛙属于雨蛙科（Hylidae），与蛙科物种的演化分歧程度较高，但我们仍鉴定出56个足够保守的基因，可通过Primer3进行非简并引物设计。本研究不仅为比较基因组学与定量基因表达分析提供了研究基础，其结果还可快速开发用于系统发育学或群体遗传学的核序列标记。