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Diapause is a state of developmental and metabolic dormancy that precedes exposure to environmental stresses. Yet, diapausing embryos are typically stress-tolerant. Evidence suggests that diapausing embryos “prepare” for stress as part of a gene expression program as they enter dormancy. Here, we investigate if diapause II embryos of the annual killifish Austrofundulus limnaeus, which can survive for hundreds of days of anoxia, can mount a transcriptomic response to anoxic insult. Bulk RNAseq was used to characterize the transcriptomes of diapause II embryos exposed to normoxia, 4 h and 24 h anoxia, and 2 h and 24 h normoxic recovery from anoxia. Differential expression and gene ontology analyses were used to probe for pathways that may mitigate survival. Transcriptional factor analysis was used to predict potential mediators of this response. Diapausing embryos exhibited a robust transcriptomic response to anoxia and recovery that returns to near baseline conditions after 24 h. Anoxia induced an upregulation of genes involved in the integrated stress response, lipid metabolism, p38mapk kinase signaling, and apoptosis. Developmental and mitochondrial genes decreased. We conclude that diapause II embryos mount a robust transcriptomic stress response when faced with anoxic insult. This response is consistent with mediating expected challenges to cellular homeostasis in anoxia.