Protocol for isolating nuclei from frozen fish heart tissue for single-cell genomic assays.

Unbiased isolation of intact single nuclei from complex tissues, such as cryopreserved hearts, for massively parallel single-cell genomic assays is particularly challenging. Here, we present a protocol for isolating high-quality nuclei from the heart tissue of Trematomus bernacchii, an Antarctic fish species with notoriously challenging tissue for nuclear extraction. We describe steps for pulverizing frozen heart tissue, library construction, and sequencing. This protocol can provide a valuable reference for isolating nuclei from other cryopreserved marine fish tissues.