Single cell RNA-sequencing reveals cellular heterogeneity and trajectories of lineage specification during murine embryonic limb development

The coordinated spatial and temporal regulation of gene expression in the murine hindlimb determines the identity of mesenchymal progenitors and the development of diversity of musculoskeletal tissues they form. Hindlimb development has historically been studied with lineage tracing of individual genes selected a priori, or at the bulk tissue level, which does not allow for the determination of single cell transcriptional programs yielding mature cell types and tissues. To identify the cellular trajectories of lineage specification during limb bud development, we used single cell mRNA sequencing (scRNA-seq) to profile the developing murine hindlimb between embryonic days (E)11.5-E18.5. We found cell type heterogeneity at all time points, and the expected cell types that form the mouse hindlimb. In addition, we used RNA fluorescence in situ hybridization (FISH) to examine the spatial locations of cell types and cell trajectories to understand the ancestral continuum of cell maturation. This data provides a resource for the transcriptional program of hindlimb development that will support future studies of musculoskeletal development and generate hypotheses for tissue regeneration. Four samples from murine embryonic hindlimb tissue at E11.5, E13.5, E15.5, and E18.5小鼠后肢内基因表达的协同时空调控，决定了间充质祖细胞（mesenchymal progenitors）的细胞身份，以及其分化形成的肌肉骨骼组织（musculoskeletal tissues）的多样性发育进程。长期以来，后肢发育的研究多采用预先筛选的单个基因的谱系示踪（lineage tracing）技术，或基于整体组织（bulk tissue）水平的分析策略，但此类方法无法解析生成成熟细胞类型与组织的单细胞转录程序（single cell transcriptional programs）。为解析肢芽（limb bud）发育过程中谱系特化的细胞轨迹，我们采用单细胞mRNA测序（single cell mRNA sequencing, scRNA-seq）技术，对胚胎期11.5天至18.5天（E11.5-E18.5）的发育中小鼠后肢进行了转录组谱分析。我们在所有采样时间点均检测到细胞类型异质性，且覆盖了构成小鼠后肢的所有预期细胞类型。此外，我们采用RNA荧光原位杂交（RNA fluorescence in situ hybridization, FISH）技术，检测了各类细胞类型及细胞轨迹的空间定位，以解析细胞成熟的祖先谱系连续体。本数据集为后肢发育的转录程序提供了宝贵的研究资源，将为后续肌肉骨骼发育相关研究提供支撑，并为组织再生领域生成可验证的科学假说。本数据集包含4份分别取自E11.5、E13.5、E15.5及E18.5的小鼠胚胎后肢组织样本。