Taketsuru Hiroaki, Kaneko Takehito
National Institute for Basic Biology, Interuniversity Bio-Backup Project, Aichi, 444-8787, Japan.
Division of Science and Engineering Graduate School of Arts and Science, Iwate University, Iwate, 020-8551, Japan; Department of Chemistry and Biological Sciences, Faculty of Science and Engineering, Iwate University, Iwate, 020-8551, Japan; Soft-Path Science and Engineering Research Center (SPERC), Iwate University, Iwate, 020-8551, Japan.
Cryobiology. 2018 Oct;84:1-3. doi: 10.1016/j.cryobiol.2018.09.002. Epub 2018 Sep 11.
Numerous genetically engineered rat strains have been produced via genome editing. Although freezing of embryos is helpful for the production and storage of these valuable strains, the tolerance to freezing of embryos varies at each developmental stage of the embryo. This study examined the tolerance to freezing of rat embryos at various developmental stages, particularly at the pronuclear stage. Embryos that had developed to the pronuclear, 2-cell, and morula stages were frozen via vitrification using ethylene glycol- and propylene glycol-based solutions. More than 90% of the embryos at all developmental stages survived after warming. The developmental rates to offspring of thawed embryos at the pronuclear, 2-cell, and morula stages were 19%, 41%, and 52%, respectively. Pronuclear stage embryos between the early and late developmental stages were then vitrified. The developmental rates to offspring of the thawed pronuclear stage embryos collected at 24, 28, and 31 h after the induction of ovulation were 17%, 21%, and 23%, respectively. These results indicated that the tolerance to vitrification of rat embryos increased with the development of embryos. The establishment of vitrification method of rat embryos at various developmental stages is helpful for improving the production and storage of valuable rat strains used for biomedical science.
通过基因组编辑已产生了许多基因工程大鼠品系。尽管胚胎冷冻有助于这些珍贵品系的生产和保存，但胚胎在每个发育阶段对冷冻的耐受性各不相同。本研究检测了大鼠胚胎在不同发育阶段，特别是原核期对冷冻的耐受性。发育至原核期、2细胞期和桑葚胚期的胚胎使用基于乙二醇和丙二醇的溶液通过玻璃化法进行冷冻。所有发育阶段的胚胎在解冻后90%以上存活。解冻后的原核期、2细胞期和桑葚胚期胚胎发育为后代的比率分别为19%、41%和52%。然后对处于原核期早期和晚期发育阶段之间的胚胎进行玻璃化冷冻。在诱导排卵后24、28和31小时收集的解冻后原核期胚胎发育为后代的比率分别为17%、21%和23%。这些结果表明，大鼠胚胎对玻璃化冷冻的耐受性随着胚胎发育而增加。建立不同发育阶段大鼠胚胎的玻璃化冷冻方法有助于改善用于生物医学的珍贵大鼠品系的生产和保存。
