Data from: A non-lethal sampling method to obtain, generate and assemble whole-blood transcriptomes from small, wild mammals

The acquisition of tissue samples from wild populations is a constant challenge in conservation biology, especially for endangered species and protected species where nonlethal sampling is the only option. Whole blood has been suggested as a nonlethal sample type that contains a high percentage of bodywide and genomewide transcripts and therefore can be used to assess the transcriptional status of an individual, and to infer a high percentage of the genome. However, only limited quantities of blood can be nonlethally sampled from small species and it is not known if enough genetic material is contained in only a few drops of blood, which represents the upper limit of sample collection for some small species. In this study, we developed a nonlethal sampling method, the laboratory protocols and a bioinformatic pipeline to sequence and assemble the whole blood transcriptome, using Illumina RNA-Seq, from wild greater mouse-eared bats (Myotis myotis). For optimal results, both ribosomal and globin RNAs must be removed before library construction. Treatment of DNase is recommended but not required enabling the use of smaller amounts of starting RNA. A large proportion of protein-coding genes (61%) in the genome were expressed in the blood transcriptome, comparable to brain (65%), kidney (63%) and liver (58%) transcriptomes, and up to 99% of the mitogenome (excluding D-loop) was recovered in the RNA-Seq data. In conclusion, this nonlethal blood sampling method provides an opportunity for a genomewide transcriptomic study of small, endangered or critically protected species, without sacrificing any individuals.

在保护生物学领域，获取野生种群的组织样本始终是一项核心难题，对于仅能采用非致死采样方式的濒危物种与受保护物种而言尤为棘手。全血作为一类非致死性样本类型，因含有大量全身及全基因组范围的转录本，被广泛提议用于评估个体的转录状态，并推断基因组的大部分序列信息。然而，针对小型物种的非致死采血采样量往往十分有限，且目前尚不明确仅几滴血液（这是部分小型物种采样的上限）中是否含有足够的遗传物质。本研究以野生大鼠耳蝠（Myotis myotis）为研究对象，基于Illumina RNA测序（Illumina RNA-Seq）技术，开发了一套用于全血转录组测序与组装的非致死采样方法、标准化实验室操作流程及生物信息学分析流程。若要获得最优测序结果，需在文库构建前同步去除核糖体RNA与珠蛋白RNA。研究建议进行DNase处理，但该步骤并非强制要求，因此可使用起始量更少的初始RNA样本。该血液转录组中表达了基因组中61%的蛋白质编码基因，这一比例与脑组织（65%）、肾脏组织（63%）及肝脏组织（58%）的转录组表达水平相当；同时，RNA测序数据可覆盖线粒体基因组（不含D-loop区）高达99%的序列。综上，该非致死采血采样方法无需牺牲任何个体，即可为小型、濒危或严格保护物种开展全基因组范围的转录组学研究提供可行途径。