In this process, tremendous incidences on the molecular and cellular levels take place, driving essential developmental activities such as lineage specifications, axis patterning, and organogenesis. Detailed understandings of the molecular mechanisms of this process, such as the transcriptome and epigenome, are critical for fundamental embryology study, management of reproduction-related diseases, and regenerative medicine. [2] Given that embryonic cells are scarce yet highly heterogeneous, analysis with the single-cell resolution is thus essential for a complete knowledge of embryonic development. However, conventional cell analysis, such as DNA microarray, quantitative realtime polymerase chain reaction (PCR), and sequencing, generally requires a sample of pooled cells and thus gauges the ensemble average, masking the cell heterogeneity among the tested sample. By performing analysis on individual cells separately, one could obtain analysis with single-cell resolution.The single-cell analysis normally includes four steps: 1) single-cell isolation and manipulation, 2) sample preparation, 3) sequencing, and 4) data analysis. [3] In principle, single-cell analysis can be performed following the same testing mechanisms as the bulk assays. However, there are a few challenges Embryonic development is a fundamental physiological process that can provide tremendous insights into stem cell biology and regenerative medicine. In this process, cell fate decision is highly heterogeneous and dynamic, and investigations at the single-cell level can greatly facilitate the understanding of the molecular roadmap of embryonic development. Rapid advances in the technology of single-cell sequencing offer a perfectly useful tool to fulfill this purpose. Despite its great promise, single-cell sequencing is highly interdisciplinary, and successful applications in specific biological contexts require a general understanding of its diversity as well as the advantage versus limitations for each of its variants. Here, the technological principles of single-cell sequencing are consolidated and its applications in the study of embryonic development are summarized. First, the technology basics are presented and the available tools for each step including cell isolation, library construction, sequencing, and data analysis are discussed. Then, the works that employed single-cell sequencing are reviewed to investigate the specific processes of embryonic development, including preimplantation, peri-implantation, gastrulation, and organogenesis. Further, insights are provided on existing challenges and future research directions.
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