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卤虫(Artemia salina)胚胎、无节幼体和绒螯蟹胚胎、蚤状幼体冷冻保存初探
刘向宇
Subtype硕士
Thesis Advisor陆仁后
1992
Degree Grantor中国科学院水生生物研究所
Place of Conferral中国科学院水生生物研究所
Degree Discipline遗传学
Keyword冷冻保护剂 冷冻 卤虫 绒螯蟹 胚胎 元节幼体 蚤状幼体
Abstract本文通过石蜡切片观察发现卤虫眠卵处于囊胚期。以甘油、DMSO、葡萄糖、蔗糖为冷冻保护剂对卤虫休虑卵及卤虫脱壳卵胚胎进行液氮冷冻保存,发现保护剂对短时期吸水发育胚胎有一定的保护作用,但对吸水充分、发育至后期的胚胎没有冷冻保护作用。双重保护剂(甘油+DMSO、葡萄糖+蔗糖)比单重保护剂保护作用强,此外,作者发现饱和NaCl溶液对卤虫各个发育时期胚胎都有非常好的冷冻保护作用,虽然对后期的保护作用较弱。本文推测卤虫胚胎冷冻损伤与胚胎吸水极其有关,后期胚胎冷冻致死可能是膜内冰晶造成损伤的结果。本文还探讨了保护剂、高渗、冷冻对绒螯蟹各发育时期的影响。发现单胞期对甘油和高渗敏感,多胞期对高渗也敏感。在本文选择的条件下,DMSO和甲醇基本无毒性。单胞期和多胞期对冷冻的忍受力最差,1/4胚体期对冷冻的忍受力最强。保护剂的加入使胚胎对冷冻的忍受力有所提高。在冷冻死亡的胚胎特别是单胞期和多胞期中发现卵黄遭受严重的损伤,这可能是绒螯蟹早期胚胎冷冻致死的一个重要原因。此外,本文还发现甘油、DMSO、葡萄糖、蔗糖对卤虫无节幼体有一定的冷冻保护作用,20%甘油保护下无节幼体冷冻至-30 ℃9-10分钟存活率达80%以上。DMSO、甲醇、甘油对中华绒螯蟹蚤状幼体也有一定的冷冻保护作用,20%甲醇保护下蚤状幼体冷冻至-24 ℃8-9分钟还有70%以上的存活率。
Other AbstractThe encysted dormant Artemis salina embryos were discovered being at their blastula stage through paraffin section. The encysted and decapsulated Artemla embryos hydrated for a short time were successfully cryopreserved in liquid nitrogen with various concentrations of glycerol, DMSO, glucose and sucrose as cryoprotectants, but cryoprotectants were found having no cryoprotection for embryos which were hydrated fully and developed to late developmental stages. Dual cryoprotectants (glycerol+DMSO, glucose+sucrose) had stronger cryoprotections than single-type cryoprotectants. Saturated NaCl solution showed a powerful eryoprotection for various developmental stages of Artemia embryos, although having a weaker cryoprotection for later developmental stages. The author suggested water in embryos related to freezing damage. That freezing was lethal to the later develop mental stages was possibly because of intramembranous ice formation. Cryoprotectant toxicity, hypertonic solution effect and freezing tolerance were determined for various developmental stages of mitten crabs. One-cell embryo was sensitive to glycerol and hypertonic solution, multicell embryo was also sensitive to hypertonic solution. DMSO and methanol had little toxicity to mitten crab embryos under the condition we used. One-cell and multicell embryos were the the most sensitive to freezing. Quarter embryo was the most tolerant to freezing. The addition of cryoprotectants to the chilled solution improved the freezing tolerance of various developmental stages. Yolk was seriously damaged especially in one-cell and multicell embryos which died after freezing. Author suggested that the damage was an important reason resulting in freezing death of early embryos. Glycerol DMSO, glucose, and sucrose were found improving tolerance of Artemia nauplii to freezing. Freezed to -30 ℃ for 9-10 minutes in 20% glycerol, more than 80% of nauplii survived. DMSO, methanol and glycerol possessed some cryoprotections for Eriocheir sinensiszoea. Freezed to -24 ℃ for 8-9 minutes in 20% methanol, more than 70% of zoea survived.
Pages56
Language中文
Document Type学位论文
Identifierhttp://ir.ihb.ac.cn/handle/342005/12854
Collection学位论文
Recommended Citation
GB/T 7714
刘向宇. 卤虫(Artemia salina)胚胎、无节幼体和绒螯蟹胚胎、蚤状幼体冷冻保存初探[D]. 中国科学院水生生物研究所. 中国科学院水生生物研究所,1992.
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