Long-term stability of RNA isolated from muscle of red seabream (Pagrus major) during ice storage

Recovering high-quality intact RNA from postmortem tissue is of major concern for gene expression studies. However, it is difficult to perform RNA extraction from aquacultured fish immediately after death, as rapid and accurate skills are needed for the procedure. The objective of this study was to quantitatively assess the integrity of total RNA extracted from muscle, liver, and digestive tract tissues of red seabream stored in ice as whole bodies, at a range of time points up to 10 days postmortem, using RNA integrity number (RIN) and quantitative PCR (qPCR). The RIN of total RNA in muscle remained over 8.0 for 5 days postmortem. The RINs in the liver and digestive tract were under 5.0 at 2 days postmortem. The mRNA levels of tissue inhibitor metalloproteinase 2 (TIMP-2) and β-actin, measured using qPCR in muscle, decreased to 87.8% at 1 day postmortem and to 45.5% at 2 days postmortem, from that at 0 days postmortem. In the liver and digestive tract, the mRNA levels were not significantly changed until 1 day postmortem. These results indicate that RNA, especially from fish muscle, can be maintained at high quality for several days postmortem solely by storing the fish body in ice.