Long-term storage of oogonia and germ-line stem cells provides an alternative to the limitations associated with cryopreserving eggs of important fish species. These cell types are less vulnerable to the stresses of freezing. Cryopreservation has enormous potential for aquaculture advancement, but protocols must be developed for each species and cell type since its success hinges on various input factors. Blue Catfish Ictalurus furcatus were selected as the test species in this study because of the need to improve fry production of Blue Catfish ♂ × Channel Catfish I. punctatus ♀ hybrids, which can be facilitated by storing oogonia in gene banks. Our objective was to develop a freezing protocol for oogonia of this species. We tested different permeating and nonpermeating cryoprotectants, concentrations of these agents, and freezing rates. We proved that all three factors influenced postthaw recovery of oogonia. Of the permeating cryoprotectants, 1.0 M dimethyl sulfoxide resulted in the most live cells with the highest viability percentages, and adding 0.2 M lactose with 10% egg yolk further improved the results. There were also specific interactions in which the effects of concentration and freezing rate varied among the cryoprotectant treatments. The most effective freezing rate was −1.0°C/min, and cell viability was reduced at −2.5°C/min and −5.0°C/min. From these results, we propose adding 1.0 M dimethyl sulfoxide with 0.2 M lactose and 10% egg yolk to cryomedia and freezing it at a rate of −1.0°C/min. By developing a cryopreservation protocol for a commonly cultured catfish, this work may guide the development of protocols for other species of interest.

中文翻译：

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为蓝鲶鱼 Oogonia 制定有效的冷冻保存方案

卵原细胞和生殖系干细胞的长期储存为重要鱼类卵的冷冻保存相关限制提供了替代方案。这些细胞类型不易受到冷冻压力的影响。冷冻保存在水产养殖发展方面具有巨大潜力，但必须为每个物种和细胞类型制定协议，因为其成功取决于各种投入因素。由于需要提高Blue Catfish ♂ × Channel Catfish I. punctatus 的鱼苗产量，因此本研究选择Blue Catfish Ictalurus furcatus作为试验种♀ 杂种，这可以通过将 oogonia 存储在基因库中来促进。我们的目标是为该物种的卵原细胞制定冷冻方案。我们测试了不同的渗透性和非渗透性冷冻保护剂、这些试剂的浓度和冷冻率。我们证明了所有三个因素都会影响卵原细胞的解冻后恢复。在渗透性冷冻保护剂中，1.0 M 二甲基亚砜导致最多活细胞的存活率最高，添加 0.2 M 乳糖和 10% 蛋黄进一步改善了结果。还有一些特定的相互作用，其中浓度和冷冻速率的影响在冷冻保护剂处理之间有所不同。最有效的冷冻速率为 -1.0°C/min，细胞活力在 -2.5°C/min 和 -5.0°C/min 时降低。根据这些结果，我们建议加 1。0 M 二甲基亚砜与 0.2 M 乳糖和 10% 蛋黄混合到低温培养基中，并以 -1.0°C/分钟的速度冷冻。通过为普遍养殖的鲶鱼制定冷冻保存协议，这项工作可以指导其他感兴趣物种的协议的制定。