High-Throughput Transcriptome Profiling of Single Nuclei and Single Synapses Using Single-Cell Total-RNA-Seq [Mm]

We developed the first droplet-based single-cell total-RNA-seq method. We refer to this platform as Multiple Annealing and Tailing-based Quantitative scRNA-seq in Droplet (MATQ-Drop). With the detection of nascent RNA species, we showed that the cell atlas of human brain samples could be effectively constructed based on nascent RNA species. Furthermore, we observed that only lncRNA species are sufficient to construct the cell atlas, suggesting that MATQ-Drop allows a large-scale identification of the cell-type-specific lncRNA species. Beyond total-RNA profiling for single nuclei, we also showed that MATQ-Drop could be used to profile the transcripts in different neuronal sub-compartments. Based on transcriptome profiling, we were able to determine different types of sub-compartments in particular, synapses. Here we have referred to the transcriptome of individual synapses as the synaptome, and we identified different subtypes of pre-synapses and post-synapses. Furthermore, between pre-synapses and post-synapses, as well as between pre-synapses and the nuclei, we were able to identify different functional enrichments. Our result unveils unprecedented new insights about gene expression in individual synapses. It also demonstrates the feasibility of using MATQ-drop to profile the transcriptome of sub-cellular compartments. MATQ-Drop single-cell total-RNA-seq of Hoechst-positive nuclei and Hoechst-negative subneuronal compartments isolated from WT and 5XFAD mouse hippocampus本团队开发了首款基于微滴的单细胞总RNA测序（single-cell total-RNA-seq）技术。我们将该技术平台命名为基于多重退火与加尾的微滴定量单细胞RNA测序（Multiple Annealing and Tailing-based Quantitative scRNA-seq in Droplet，简称MATQ-Drop）。借助对新生RNA分子的检测，我们证实可基于新生RNA分子高效构建人脑样本的细胞图谱。此外，我们发现仅需长链非编码RNA（long non-coding RNA，lncRNA）分子即可构建细胞图谱，这表明MATQ-Drop可实现细胞类型特异性长链非编码RNA分子的大规模鉴定。除针对单细胞核开展总RNA谱分析外，我们还证实MATQ-Drop可用于分析不同神经元亚区室的转录本。基于转录组谱分析，我们可区分不同类型的亚区室，尤其是突触。本研究将单个突触的转录组命名为“突触组（synaptome）”，并鉴定出突触前与突触后神经元的不同亚型。此外，我们还在突触前与突触后神经元之间，以及突触前神经元与细胞核之间，鉴定出不同的功能富集特征。本研究结果为单个突触的基因表达研究提供了前所未有的新认知，同时也证实了利用MATQ-Drop分析亚细胞区室转录组的可行性。本数据集包含从野生型（wild type，WT）与5XFAD小鼠海马体中分离的Hoechst阳性细胞核及Hoechst阴性神经元亚区室的MATQ-Drop单细胞总RNA测序数据。