Smith College, Department of Biological Sciences, Northampton, MA, USA.
Smith College, Department of Biological Sciences, Northampton, MA, USA; University of Massachusetts Amherst, Program in Organismic and Evolutionary Biology, Amherst, MA, USA.
Eur J Protistol. 2021 Oct;81:125840. doi: 10.1016/j.ejop.2021.125840. Epub 2021 Sep 4.
Though representing a major component of eukaryotic biodiversity, many microbial eukaryotes remain poorly studied, including the focus of the present work, testate amoebae of the order Arcellinida (Amoebozoa) and non-model lineages of ciliates (Alveolata). In particular, knowledge of genome structures and changes in genome content over the often-complex life cycles of these lineages remains enigmatic. However, the limited available knowledge suggests that microbial eukaryotes have the potential to challenge our textbook views on eukaryotic genomes and genome evolution. In this study, we developed protocols for DAPI (4',6-diamidino-2-phenylindole) staining of Arcellinida nuclei and adapted protocols for ciliates. In addition, image analysis software was used to estimate the DNA content in the nuclei of Arcellinida and ciliates, and the measurements of target organisms were compared to those of well-known model organisms.The results demonstrate that the methods we have developed for nuclear staining in these lineages are effective and can be applied to other microbial eukaryotic groups by adjusting certain stages in the protocols.
尽管真核生物多样性的主要组成部分，但许多微生物真核生物仍未得到充分研究，包括本工作的重点，即壳质目（Amoebozoa）的有壳变形虫和纤毛类的非模式谱系（Alveolata）。特别是，这些谱系的复杂生命周期中，有关基因组结构和基因组内容变化的知识仍然是个谜。然而，有限的现有知识表明，微生物真核生物有可能挑战我们关于真核生物基因组和基因组进化的教科书观点。在这项研究中，我们开发了用于壳质目细胞核 DAPI（4'，6-二脒基-2-苯基吲哚）染色的方案，并对纤毛虫的方案进行了改编。此外，使用图像分析软件来估计壳质目和纤毛虫细胞核中的 DNA 含量，并将目标生物的测量值与知名模型生物的测量值进行比较。结果表明，我们为这些谱系中的核染色开发的方法是有效的，可以通过调整方案中的某些阶段应用于其他微生物真核生物群体。
