This study was carried out to test how sperm cryopreservation affected nuclear DNA stability and whether progeny development was modified when eggs were fertilized with cryopreserved spermatozoa. The "comet assay" (alkaline single-cell gel electrophoresis assay) was adapted to trout spermatozoa to estimate DNA stability as measured by alkali-induced DNA strand break formation. Because trout eggs develop in water after fertilization (oviparous species) and that eggshell is easy to clear up after fixative treatment, progeny development was assessed from the blastodisc flattening stage of the embryos to the first feeding stage of the hatched fries by direct observation. All parameters under study were analyzed on each sperm and comparisons between parameters were made using paired data. Freeze-thawing of sperm slightly but significantly increased the percentage of nuclei showing altered DNA after comet assay. This increase was correlated to the decrease in fertilization rates of sperm, but the absolute percentage of altered nuclei was not predictive of the absolute fertilization ability of sperm. Assessment of progeny development showed that survival rate and abnormality rate obtained after fertilization with cryopreserved sperm were not different from those obtained with fresh sperm. It is concluded that trout sperm cryopreservation only slightly affected sperm DNA stability and that the use of cryopreserved spermatozoa did not impair offspring survival and quality.

译文

进行这项研究是为了测试精子冷冻保存如何影响核DNA的稳定性,以及当用冷冻保存的精子使卵受精时是否改变了后代的发育。“彗星测定” (碱性单细胞凝胶电泳测定) 适用于鳟鱼精子,以估算通过碱诱导的DNA链断裂形成测得的DNA稳定性。由于鳟鱼卵在受精后 (卵生物种) 在水中发育,并且在固定处理后蛋壳易于清除,因此通过直接观察评估了从胚胎的胚盘扁平阶段到孵化的薯条的第一个饲养阶段的后代发育。对每个精子分析了所有研究参数,并使用配对数据进行了参数之间的比较。彗星分析后,精子的冻融略有增加,但显着增加了显示DNA改变的核百分比。这种增加与精子受精率的降低有关,但是细胞核改变的绝对百分比不能预测精子的绝对受精能力。对后代发育的评估表明,冷冻保存的精子受精后的存活率和异常率与新鲜精子受精后的存活率和异常率没有差异。结论是,鳟鱼精子冷冻保存仅轻微影响精子DNA的稳定性,并且使用冷冻保存的精子不会损害后代的存活率和质量。

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