Abstract: Sperm cryopreservation is a key technique for germplasm resource conservation in aquatic animals. However, the damage and response mechanisms of black tiger shrimp (Penaeus monodon) sperm after cryopresercation remain unclear. In this study, we first analyzed the impact of cryopreservation on sperm motility. Subsequently, we employed high-throughput sequencing to systematically compare the expression profiles of messenger RNAs (mRNAs) and long non-coding RNAs (LncRNAs) between fresh and cryopreserved sperm. The results show that both sperm motility and fertilization rate were significantly lower in the cryopreserved group compared with the fresh group. Transcriptomic analysis identified a total of 267 differentially expressed mRNAs (6 up-regulated and 261 down-regulated) and 884 differentially expressed LncRNAs (52 up-regulated and 832 down-regulated). Enrichment analysis reveals that the differentially expressed mRNAs were primarily involved in biological processes such as RNA-dependent DNA replication, biological regulation, intracellular signal transduction, ATP binding, and membrane integral components, and were significantly enriched in pathways related to lipid metabolism, signal transduction, and cytoskeleton. The target genes regulated by LncRNAs were significantly enriched in biological processes associated with cell membrane, signal transduction, protein degradation, and energy metabolism. This study clarifies the potential synergistic role of LncRNAs and mRNAs in the cryostress response, providing new theoretical insights for further deciphering the molecular mechanisms of sperm cryodamage and optimizing cryopreservation strategies.