Single‐Cell Sequencing: A Sharp Sword for Deciphering Biological Innovations and Evolutionary Adaptations in Lower Vertebrates and Invertebrates

ABSTRACT Single‐cell sequencing (SCS) technology has revolutionized the study of cellular heterogeneity by enabling the exploration of biological processes at high resolution. While traditionally applied to mammalian models, recent advancements have extended SCS to ecologically and evolutionarily crucial lower vertebrates (particularly fish) and diverse invertebrates (including Urochordata, Echinodermata, Arthropoda, Mollusca, and Coelenterata). These non‐mammalian organisms exhibit unique cellular and molecular adaptations, serving as pivotal models for developmental biology, immune mechanisms, hematopoiesis, and environmental adaptation, yet remain underrepresented in genomics. This review highlights how SCS has been instrumental in uncovering novel cell types, defining intricate developmental trajectories, revealing unprecedented complexity in hematopoiesis, and elucidating diverse immune landscapes (including cell heterogeneity, functional specialization, pathogen responses, and tissue‐specific adaptations) in fish. Furthermore, it details the construction of foundational cell atlases across fish and invertebrate species and showcases SCS applications in mapping invertebrate development, characterizing cellular heterogeneity under varying conditions, and revealing unique environmental adaptations. Cross‐species SCS analyses provide powerful tools for tracing the evolution of critical systems, such as immunity. While addressing challenges like incomplete reference genomes and technical complexity, we discuss SCS's transformative potential in aquaculture and biotechnology, emphasizing future directions in multi‐omics integration and computational tool development to decipher biological innovations and evolutionary adaptations.