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Abstract

The transformative events during early organismal development lay the foundation for body formation and long-term phenotype. The rapid progression of events and the limited material available present major barriers to studying these earliest stages of development. Herein, we report an operationally simple RNA sequencing approach for high-resolution, time-sensitive transcriptome analysis in early (≤3 h) Drosophila embryos. This method does not require embryo staging but relies on single-embryo RNA sequencing and transcriptome ordering along a developmental trajectory (pseudo-time). The resulting high-resolution, time-sensitive mRNA expression profiles reveal the exact onset of transcription and degradation for thousands of transcripts. Further, using sex-specific transcription signatures, embryos can be sexed directly, eliminating the need for Y chromosome genotyping and revealing patterns of sex-biased transcription from the beginning of zygotic transcription. Our data provide an unparalleled resolution of gene expression during early development and enhance the current understanding of early transcriptional processes.