Lin C, Spikings E, Zhang T, Rawson D M
LIRANS Institute of Research in the Applied Natural Sciences, University of Bedfordshire, Bedfordshire, UK.
Cryobiology. 2009 Aug;59(1):42-7. doi: 10.1016/j.cryobiol.2009.04.007. Epub 2009 May 3.
Cryopreservation is now common practice in the fields of aquaculture, conservation and biomedicine. However, there is a lack of information on the effect of chilling and cryopreservation at the molecular level. In the present study, we used real-time RT-PCR analysis to determine the effect of chilling and cryopreservation on expression of Pax2a, Pax2b, Pax5 and Pax8 which constitute one subgroup of the Pax gene family. As intact embryos of zebrafish have not yet been successfully cryopreserved, we have used two alternatives: chilling of intact embryos and cryopreservation of isolated blastomeres. Cryopreservation was found to affect the normal pattern of gene expression in zebrafish embryonic blastomeres. The trends, profile changes, in expression of Pax2a and Pax5 occurred to a lesser extent in frozen-thawed blastomeres than in fresh blastomeres whilst the opposite was true for Pax8. The trends in expression of Pax2b were delayed in frozen-thawed blastomeres compared to fresh blastomeres. Cryopreservation can therefore disrupt normal gene expression patterns in zebrafish embryonic blastomeres which could have a detrimental effect on embryo development.
冷冻保存如今在水产养殖、保护生物学和生物医学领域已是常见做法。然而，在分子水平上，关于冷休克和冷冻保存的影响仍缺乏相关信息。在本研究中，我们使用实时逆转录聚合酶链反应（RT-PCR）分析来确定冷休克和冷冻保存对构成Pax基因家族一个亚组的Pax2a、Pax2b、Pax5和Pax8表达的影响。由于斑马鱼完整胚胎尚未成功实现冷冻保存，我们采用了两种替代方法：完整胚胎的冷休克处理和分离的卵裂球的冷冻保存。研究发现，冷冻保存会影响斑马鱼胚胎卵裂球中基因表达的正常模式。与新鲜卵裂球相比，Pax2a和Pax5在冻融卵裂球中的表达趋势、图谱变化程度较小，而Pax8的情况则相反。与新鲜卵裂球相比，Pax2b在冻融卵裂球中的表达趋势出现延迟。因此，冷冻保存可能会扰乱斑马鱼胚胎卵裂球中的正常基因表达模式，这可能会对胚胎发育产生不利影响。
