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Cell cycle synchronisation of Trypanosoma brucei by centrifugal counter-flow elutriation reveals the timing of nuclear and kinetoplast DNA replication

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@article{0f65ca24f079414eb46272b70f040255,
title = "Cell cycle synchronisation of Trypanosoma brucei by centrifugal counter-flow elutriation reveals the timing of nuclear and kinetoplast DNA replication",
abstract = "We report an optimised centrifugal counter-flow elutriation protocol for the rapid and direct isolation of G1 cell cycle synchronised populations of both the procyclic and bloodstream form stages of Trypanosoma brucei that yields viable and proliferative cells. The high quality of the synchronisation achieved can be judged by the uniform DNA content, narrow size distribution, synchronous division, and the maintenance of synchronicity into subsequent cell cycles. We show that early-eluting fractions represent different G1 subpopulations that progress through the cell cycle with distinct temporal profiles post-elutriation, as exemplified by the observation of the maturation of a second flagellar basal body in late G1 phase, DNA replication in S phase, and dimethylation of histone H3 in mitosis/cytokinesis. We use our temporal observations to construct a revised model of the relative timing and duration of the nuclear and kinetoplast cell cycle that differs from the current model.",
keywords = "DNA replication, Isolation, separation and purification, Parasite biology",
author = "Corinna Benz and Frank Dondelinger and McKean, {Paul Gerard} and Urbaniak, {Michael Daniel}",
year = "2017",
month = dec,
day = "14",
doi = "10.1038/s41598-017-17779-z",
language = "English",
volume = "7",
journal = "Scientific Reports",
issn = "2045-2322",
publisher = "Nature Publishing Group",

}

RIS

TY - JOUR

T1 - Cell cycle synchronisation of Trypanosoma brucei by centrifugal counter-flow elutriation reveals the timing of nuclear and kinetoplast DNA replication

AU - Benz, Corinna

AU - Dondelinger, Frank

AU - McKean, Paul Gerard

AU - Urbaniak, Michael Daniel

PY - 2017/12/14

Y1 - 2017/12/14

N2 - We report an optimised centrifugal counter-flow elutriation protocol for the rapid and direct isolation of G1 cell cycle synchronised populations of both the procyclic and bloodstream form stages of Trypanosoma brucei that yields viable and proliferative cells. The high quality of the synchronisation achieved can be judged by the uniform DNA content, narrow size distribution, synchronous division, and the maintenance of synchronicity into subsequent cell cycles. We show that early-eluting fractions represent different G1 subpopulations that progress through the cell cycle with distinct temporal profiles post-elutriation, as exemplified by the observation of the maturation of a second flagellar basal body in late G1 phase, DNA replication in S phase, and dimethylation of histone H3 in mitosis/cytokinesis. We use our temporal observations to construct a revised model of the relative timing and duration of the nuclear and kinetoplast cell cycle that differs from the current model.

AB - We report an optimised centrifugal counter-flow elutriation protocol for the rapid and direct isolation of G1 cell cycle synchronised populations of both the procyclic and bloodstream form stages of Trypanosoma brucei that yields viable and proliferative cells. The high quality of the synchronisation achieved can be judged by the uniform DNA content, narrow size distribution, synchronous division, and the maintenance of synchronicity into subsequent cell cycles. We show that early-eluting fractions represent different G1 subpopulations that progress through the cell cycle with distinct temporal profiles post-elutriation, as exemplified by the observation of the maturation of a second flagellar basal body in late G1 phase, DNA replication in S phase, and dimethylation of histone H3 in mitosis/cytokinesis. We use our temporal observations to construct a revised model of the relative timing and duration of the nuclear and kinetoplast cell cycle that differs from the current model.

KW - DNA replication

KW - Isolation, separation and purification

KW - Parasite biology

U2 - 10.1038/s41598-017-17779-z

DO - 10.1038/s41598-017-17779-z

M3 - Journal article

VL - 7

JO - Scientific Reports

JF - Scientific Reports

SN - 2045-2322

M1 - 17599

ER -