Instituto de Biologia Experimental e Tecnológica (IBET)/Instituto de Tecnologia Química e Biológica-Universidade Nova de Lisboa (ITQB-UNL), Av. República (EAN) 2784-505 Oeiras, Portugal.
J Biotechnol. 2009 Sep 25;143(4):288-95. doi: 10.1016/j.jbiotec.2009.08.001. Epub 2009 Aug 7.
An approach combining morphological profiling and flow cytometric analysis was used to assess genetic stability during the several steps of somatic embryogenesis in Pinus pinaster. Embryogenic cell lines of P. pinaster were established from immature zygotic embryos excised from seeds obtained from open-pollinated trees. During the maturation stage, phenotype of somatic embryos was characterized as being either normal or abnormal. Based upon the prevalent morphological traits, different types of abnormal embryos underwent further classification and quantification. Nuclear DNA content of maritime pine using the zygotic embryos was estimated to be 57.04 pg/2C, using propidium iodide flow cytometry. According to the same methodology, no significant differences (P< or =0.01) in DNA ploidy were detected among the most frequently observed abnormal phenotypes, embryogenic cell lines, zygotic and normal somatic embryos, and somatic embryogenesis-derived plantlets. Although the differences in DNA ploidy level do not exclude the occurrence of a low level of aneuploidy, the results obtained point to the absence of major changes in ploidy level during the somatic embryogenesis process of this economically important species. Therefore, our primary goal of true-to-typeness was assured at this level.
采用形态分析和流式细胞术分析相结合的方法，评估了在欧洲赤松体胚发生的几个步骤中的遗传稳定性。从来自开放授粉树木的种子中取出的未成熟合子胚中建立了欧洲赤松的胚性细胞系。在成熟阶段，体胚的表型特征表现为正常或异常。基于流行的形态特征，对不同类型的异常胚胎进行了进一步的分类和定量。使用碘化丙啶流式细胞术估计使用海松合子胚的欧洲赤松的核 DNA 含量为 57.04 pg/2C。根据相同的方法，在最常观察到的异常表型、胚性细胞系、合子和正常体胚以及体细胞胚发生衍生的植物中，未检测到 DNA 倍性的显著差异（P<或=0.01）。尽管 DNA 倍性水平的差异不能排除低水平非整倍体的发生，但获得的结果表明，在该经济上重要的物种的体细胞胚胎发生过程中，倍性水平没有发生重大变化。因此，我们在这个水平上确保了真正的典型性。
