中华绒螯蟹胚胎的玻璃化冷冻保存

研究了中华绒螯蟹(Eriocheir sinensis)4个不同发育时期胚胎对A号玻璃化液的耐受性和玻璃化冷冻保存。结果表明,不同时期的胚胎对玻璃化液的耐受能力不同,其中卵裂期胚胎对玻璃化液的耐受能力较差(20~30 min),前无节幼体期和原溞状幼体期胚胎在玻璃化液中的适应时间较长(40~60 min);随着平衡时间的延长,中华绒螯蟹各个时期的胚胎成活率逐渐下降。中华绒螯蟹前无节幼体期胚胎在A号玻璃化液中平衡40 min,0.25mol/L的蔗糖分别洗脱5、10、15、20 min后,胚胎成活率无显著性差异(P>0.05)。前无节幼体期胚胎在A号玻璃化液中平衡40 min,在–196℃冷冻40 min,快速解冻后用0.25 mol/L的蔗糖洗脱10 min,有8个胚胎成活,成活率为(9.3±2.5)%,胚胎培养至第4天死亡;原溞状幼体期胚胎在A号玻璃化液中平衡40 min,在–196℃冷冻35 min,经相同浓度的蔗糖洗脱相同的时间,有7个胚胎成活,成活率(11.3±3.6)%,培养至第6天时,1个胚胎孵化出膜,出膜胚胎成活1d后死亡。

Cryopreservation of Eriocheir sinensis embryos by vitrification

to code A vitrifying solution and vitrification cryopreservation at four stages of embryonic development. Resistance to the vitrifying solution varied during embryonic development, being lowest among cleavage stage embryos (2060 min). Embryo survival decreased as the equilibration time in code A vitrifying solution increased. The survival of pre-nauplius stage embryos was not different when embryos were equilibrated for 40 min in code A vitrifying solution then eluted for 5, 10, 15, or 20 min using 0.25 mol/L sucrose. We measured survival of pre-nauplius stage embryos that were equilibrated for 40 min in code A vitrifying solution, frozen for 40 min at , then thawed rapidly and eluted for 10 min with sucrose. Eight embryos (this process, but died on 4 d after undergoing the freeze/thaw treatment. The experiment was repeated using protozoea stage embryos, although each embryo was only frozen for 35 min at % survived and one embryo hatched on the sixth day post freeze/thaw. This individual died 1 d after hatch.