Dry artificial seeds through induction of desiccation tolerance of somatic embryos by abscisic acid and study of mechanism of the desiccation tolerance

人工种子技术可能在植物繁殖系统中提供潜在的益处。如果体细胞胚在含水量降至真种子水平后仍能保持活力，则可作为种质资源储存方法和植物发育生物学研究的模式系统以及人工种子的利用。本研究旨在探讨外源外源ABA对芸苔属作物小孢子胚脱水耐性的诱导作用，并对其脱水耐性胚的蛋白和基因进行分析。干燥处理后，胚的相对含水量下降到10%左右。ABA诱导了胚的干燥耐受性。这些现象受胚大小和ABA浓度的影响，但与基因型无关。10^<-4>-10^<-5>M ABA处理的干子叶胚植株转化率为41-53%。相比之下，当不添加ABA时，几乎所有胚胎在干燥后都失去了生存能力。即使添加ABA，球形胚也不能获得干燥耐受性。虽然高渗剂也能诱导胚的耐干性，但植株转化率（1.3 ~ 8.3%）低于ABA处理。干燥胚在室温条件下保存6个月后仍能保持植株转化能力。从干燥胚中获得的植株在形态上与未干燥胚无明显差异。蛋白质的SDS-PAGE和western blot分析显示，ABA处理的胚胎积累了种子贮藏蛋白。二维凝胶分析表明，在ABA处理过程中合成了一些新的多肽，这些多肽可能与获得干燥耐受性有关。以Lea基因（pLEA76）为探针，筛选了ABA处理胚胎中mRNA的cDNA文库。选择了6个cDNA克隆。现在我们来描述这些cDNA克隆。少

Artificial seed technology may provide potential benefits in plant propagation systems. If somatic embryos could maintain their viability after their water content was decreased to the level of true seeds, they were used as a method of germplasm storage and a model system of study of developmental plant biology as well as a utilization of artificial seed. The objective of this study was to investigate induction of desiccation tolerance in microspore-derived embryos of Brassica crops by exogenous application of ABA and to analyze the proteins and genes of the desiccation tolerant embryos.After desiccation treatment, the relative water content of embryos had decreased to about 10%. Desiccation tolerance of the embryos was induced by ABA. These phenomena were influenced by the size of embryos and the concentration of ABA, but not dependent on genotypes. The frequency of plant conversion of dry cotyledonary embryos which were treated with 10^<-4>-10^<-5>M ABA was 41-53%. In contrast, almos … More t all embryos lost their viability after desiccation, when they were not treated with ABA. Globular embryos could not acquire desiccation tolerance, even if they were treated with ABA. Though desiccation tolerance of the embryos was also induced by high osmoticum, the frequency of plant conversion (1.3-8.3%) was lower than that treated with ABA. The dry embryos maintained their plant conversion ability after six months of storage under room temperature conditions. The plantlets recovered from dry embryos were not morphologically different from those from non-desiccated ones.SDS-PAGE and western blot analysis of proteins revealed that the embryos treated with ABA accumulated seed storage proteins. And two dimensional gel analysis of the embryo proteins indicated that several new polypeptides were synthesized during ABA treatment and they may be associated with the acquisition of desiccation tolerance. A cDNA library representing mRNA's present in embryos treated with ABA was screened using Lea gene (pLEA76) as a probe. Six cDNA clones were selected. Now we are characterizing these cDNA clones. Less