Nanopore-Based Sequencing of the Full-Length Transcriptome of Male and Female Cleavage-Stage Embryos of the Chinese Mitten Crab ()

The cleavage stage plays a crucial role in embryo development, characterized by a swift surge in cell proliferation alongside the accurate genetic material transmission to offspring. To delve into the characteristics of sex development during the cleavage stage of embryos, we generated the full-length transcriptome of male and female cleavage-stage embryos using Oxford Nanopore Technologies (ONT). Notably, this investigation represents the first sequencing effort distinguishing between genders in embryos. In the transcriptome structure analysis, male and female cleavage-stage embryos, while not clustered, exhibited a comparable frequency of alternative splicing (AS) occurrences. We also successfully identified 2875 transcription factors (TFs). The quantitative analysis showed the top 150 genes, in which the highly expressed genes in male embryos predominantly related to protein synthesis and metabolism. Further investigation unveiled 500 differentially expressed genes (DEGs), of which 7 male-biased ribosomal protein genes (RPGs) were particularly noteworthy and further confirmed. These analyses suggest that there may be a more active protein synthesis process in male cleavage-stage embryos. Furthermore, among the 2875 identified TFs, we predicted that 18 TFs could regulate the differentially expressed RPGs, with most TFs belonging to the zf-C2H2 and Homeobox families, which are crucial for embryonic development. During the cleavage stage of , the differential RPGs between genders were intricately linked to energy metabolism. We proposed that these RPGs exert regulatory effects on gene expression in , thereby regulating the difference of development between male and females. Our research sheds light on the developmental mechanisms of during the embryo stage and establishes a groundwork for a deeper understanding of sex development in . The results also provide comprehensive full-length transcriptome data for future gene expression and genetic studies in .