Valladares Silvia, Toribio Mariano, Celestino Cristina, Vieitez Ana M
Instituto de Investigaciones Agrobiologicas de Galicia, CSIC, Apdo. 122, 15080 Santiago de Compostela, Spain.
Cryo Letters. 2004 May-Jun;25(3):177-86.
Recent progress in somatic embryogenesis from selected mature trees of Quercus suber, has led to a demand for maintenance of a large number of selected embryogenic lines. To facilitate the management of this material a protocol for the long-term storage of this germplasm should be defined. This study reports on the use of a simple vitrification procedure for the successful cryopreservation of three cork oak embryogenic lines. High embryo recovery levels (88-93 percent) were obtained by first preculturing 2-4 mg clumps of two or three globular embryos on semisolid medium containing 0.3 M sucrose for three days, followed by incubation in PVS2 vitrification solution at 0 degree C for 60 min before direct immersion in liquid nitrogen. The mean number of embryos produced per explant was significantly greater for cryostored embryos than for untreated stock cultures, but the productivity of the latter was recovered in subsequent subcultures of the material produced by cryostored embryos. The germination and plant regeneration rates achieved by cultures derived from cryostored embryos, around 60 percent, were similar to those of non-cryopreserved stock cultures.
从选定的成年栓皮栎树上进行体细胞胚胎发生的最新进展，引发了对大量选定胚性系进行保存的需求。为便于管理这些材料，应制定该种质长期保存的方案。本研究报告了使用一种简单的玻璃化程序成功冷冻保存三个栓皮栎胚性系的情况。通过先将两到三个球形胚的2-4毫克团块在含有0.3M蔗糖的半固体培养基上预培养三天，然后在0℃的PVS2玻璃化溶液中孵育60分钟，再直接浸入液氮中，获得了较高的胚胎恢复率（88-93%）。冷冻保存的胚胎每个外植体产生的平均胚数显著高于未处理的原种培养物，但后者的生产力在冷冻保存胚胎产生的材料的后续继代培养中得以恢复。冷冻保存胚胎的培养物实现的发芽率和植株再生率约为60%，与未冷冻保存的原种培养物相似。
