The thawed spermatozoa do not need to be motile to achieve fertilization and support embryo development after application of ICSI, Accordingly, many simple spermatozoa preservation methods were explored. In the present study, two simple methods without cryoprotectants were investigated. We compared the fertilization and embryo development ability of sperm preserved in - 20 ℃ and -196 ℃, Moreover. DNA methylation, an important epigenetic modification, was detected in embryos of two methods. The results showed both two methods did not change the pattern of DNA methylation. Sperm freezing in - 196 ℃ compromised its fertilization and embryo development capacity which could be rescued by oocytes activation. Fertilization and embryo development capacity of sperm preserved in - 20 ℃ decreased when preserved for six months but maintained substantial fertilization and embryo development ability at one month. In conclusion, the - 20 ℃ method is simple, effective and safe.%小鼠卵胞浆内单精子注射(ICSI)的开展使解冻后的精子无需具有活动能力就可以使卵子受精和发育,因此多种简单的精子冻存方法被开发.从复苏后精子受精和支持胚胎发育的能力,以及DNA甲基化方面研究比较无冷冻保护剂的条件下-20℃和-196℃冻存的两种方法.结果表明,两种方法都不改变胚胎DNA甲基化状态,-196℃保存的精子的受精率和胚胎发育率降低,但是通过卵子激活可以修复受精和胚胎发育的能力.-20℃保存精子1个月不影响受精和胚胎发育能力,冻存6个月降低精子的受精和胚胎发育潜能.因此,-20℃短期保存精子是一种简单、有效且安全的方法.
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