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DAPI staining and DNA content estimation of nuclei in uncultivable microbial eukaryotes (Arcellinida and Ciliates).

ABSTRACT: 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.

SUBMITTER: Munyenyembe K

PROVIDER: S-EPMC8699166 | biostudies-literature | 2021 Oct

REPOSITORIES: biostudies-literature

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DAPI staining and DNA content estimation of nuclei in uncultivable microbial eukaryotes (Arcellinida and Ciliates).

Munyenyembe Ketty K Timmons Caitlin C Weiner Agnes K M AKM Katz Laura A LA Yan Ying Y

European journal of protistology 20210904

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 ...[more]

PMID: 34717075

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Project description:BackgroundTubulins are major components of the eukaryotic cytoskeletons that are crucial in many cellular processes. Ciliated protists comprise one of the oldest eukaryotic lineages possessing cilia over their cell surface and assembling many diverse microtubular structures. As such, ciliates are excellent model organisms to clarify the origin and evolution of tubulins in the early stages of eukaryote evolution. Nonetheless, the evolutionary history of the tubulin subfamilies within and among ciliate classes is unclear.ResultsWe analyzed the evolutionary pattern of ciliate tubulin gene family based on genomes/transcriptomes of 60 species covering 10 ciliate classes. Results showed: (1) Six tubulin subfamilies (α_Tub, β_Tub, γ_Tub, δ_Tub, ε_Tub, and ζ_Tub) originated from the last eukaryotic common ancestor (LECA) were observed within ciliates. Among them, α_Tub, β_Tub, and γ_Tub were present in all ciliate species, while δ_Tub, ε_Tub, and ζ_Tub might be independently lost in some species. (2) The evolutionary history of the tubulin subfamilies varied. Evolutionary history of ciliate γ_Tub, δ_Tub, ε_Tub, and ζ_Tub showed a certain degree of consistency with the phylogeny of species after the divergence of ciliate classes, while the evolutionary history of ciliate α_Tub and β_Tub varied among different classes. (3) Ciliate α- and β-tubulin isoforms could be classified into an "ancestral group" present in LECA and a "divergent group" containing only ciliate sequences. Alveolata-specific expansion events probably occurred within the "ancestral group" of α_Tub and β_Tub. The "divergent group" might be important for ciliate morphological differentiation and wide environmental adaptability. (4) Expansion events of the tubulin gene family appeared to be consistent with whole genome duplication (WGD) events in some degree. More Paramecium-specific tubulin expansions were detected than Tetrahymena-specific ones. Compared to other Paramecium species, the Paramecium aurelia complex underwent a more recent WGD which might have experienced more tubulin expansion events.ConclusionsEvolutionary history among different tubulin gene subfamilies seemed to vary within ciliated protists. And the complex evolutionary patterns of tubulins among different ciliate classes might drive functional diversification. Our investigation provided meaningful information for understanding the evolution of tubulin gene family in the early stages of eukaryote evolution.

Dataset Information

DAPI staining and DNA content estimation of nuclei in uncultivable microbial eukaryotes (Arcellinida and Ciliates).

Publications

DAPI staining and DNA content estimation of nuclei in uncultivable microbial eukaryotes (Arcellinida and Ciliates).

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