<p indent=0mm>Fresh Goji berry has become more and more popular in recent years with consumers’ increasing demand for nutritional, healthy, and natural food. However, freshly harvested Goji berry is highly perishable, deteriorating and quickly losing its commercial value in <sc>1–2 d</sc> at room temperature due to its unique structural and biological characteristics (tender, hollow with thin pericarp and high water content). In order to optimize the storage conditions for fresh Goji berry and better retain its nutritional components, Goji berry was stored at 4 and −4°C for up to <sc>21 d.</sc> The changes in fruit appearance, the evolution of the contents of zeaxanthin, β-carotene, and zeaxanthin dipalmitate, and the expressions of genes related to the synthesis, storage, and degradation of carotenoids and those related to the metabolism of amino acids were investigated in the present study. The results showed that the contents of zeaxanthin, β-carotene, and zeaxanthin dipalmitate were relatively higher in Goji berry stored at 4°C for <sc>7 d</sc> or at –4°C for <sc>21 d.</sc> As the most characteristic carotenoid in Goji berry, zeaxanthin dipalmitate was 42.14% higher at –4°C than at 4°C at the end of storage. The carotenoid synthesis genes were expressed at higher levels, while the carotenoid storage protein genes, <italic>LbHSP21</italic> and <italic>LbOR2</italic>, were expressed at significantly lower levels at 4°C. At the same time, the carotenoid cleavage enzyme genes,<italic> LbNCED6</italic>, <italic>LbCCD1</italic>, and <italic>LbCCD4</italic>, were expressed at significantly higher levels in Goji berry stored at 4°C. In particular, the expression level of <italic>LbCCD4</italic> on day 7 was 70 and 30 times higher than that on day 0 at 4 and –4°C, respectively, which may explain the lower carotenoids contents at 4°C by the end of storage. The expressions of proline synthesis genes, <italic>LbP5CS</italic> and <italic>LbOAT</italic>, increased, while that of proline degradation gene <italic>LbPRODH</italic> declined during storage. The expression level of <italic>LbOAT</italic> was significantly higher, while that of <italic>LbProDH</italic> was significantly lower at 4 than at –4°C (<italic>P</italic>< 0.01), which might facilitate the accumulation of proline in Goji berry stored at 4°C. At the same time, the continuously higher expression of aspartate kinase gene <italic>LbAK</italic> was conducive to the synthesis and accumulation of asparagine. The expression of <italic>LbC4H1</italic> increased gradually during storage, which was consistent with the changing trend of phenylalanine content. In conclusion, the contents of carotenoids and amino acids in Goji berry changed dynamically during postharvest storage, which were not only regulated by the internal gene expression but also affected by the external environment such as temperature. The results of the present study showed that Goji berry stored at –4°C had higher contents of carotenoids, a lower content of proline, and therefore, a better nutritional quality with postponed senescence and deterioration. The significantly higher expression of <italic>LbCDD4</italic> and lower expressions of carotenoid storage protein genes may be the main reason for the lower carotenoids contents at 4°C compared with at –4°C. The presented study preliminarily elucidated the possible molecular mechanism of why –4°C storage is more conducive to the accumulation of carotenoids and the delay of senescence and deterioration in Goji berry. This study provides new insights for the preservation and circulation of fresh Goji berry in the supply chain.