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McCulloch, Bryn (2021) DNA replication in growth conditions that mimic the natural habitat of Haloferax volcanii. MRes thesis, University of Nottingham.

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Abstract

The initial aim of the project was to assess origin-independent replication in Haloferax volcanii (Hfx. volcanii). DNA replication is initiated at specific sites on the chromosome called origins. Origins are assumed to be an essential feature of all cells, because they serve as binding sites for proteins that recruit the DNA replication machinery. In work published by Hawkins et al, (2013), it was demonstrated that mutants of Hfx. volcanii lacking all replication origins are viable; in fact, they grow faster than the wild-type and have no obvious cellular defects. By contrast, deletion of origins from Eukaryotes and Bacteria leads to cell death or profound growth defects.

The question addressed in this project was whether the accelerated growth of Hfx. volcanii cells in the absence of replication originsis due to an artefact created by rich laboratory media conditions. This may explain why replication origins have not been eliminated by natural selection, as in the natural habitat of Hfx. volcanii, the wild-type strain would have an evolutionary advantage. To test this, a growth competition assay was modified to use fluorescent proteins and flow cytometry. It was predicted that in low nutrient media, the growth advantage of origin-deleted mutants will be minimised or eliminated, as these phenotypes are not witnessed in a natural environment.

However, due to the outbreak of the COVID-19 pandemic, the project was altered to examine which factors are required for an organism to replicate without origins. A bioinformatic approach was chosen, adapting previously created tools to better fit a large data set and to predict the ability of 85 species to survive without origins. The bioinformatic pipeline involved a principal component analysis, which would take into account for any given species their respective nucleotide skew indices, spectral ratios, information gene linkage, co-orientation of core genes with DNA replication, and types of DNA polymerase genes located near origins. The results suggested several new candidate species for further experimentation and potential directions for improvement of the origin independent replication prediction tool.

Item Type:	Thesis (University of Nottingham only) (MRes)
Supervisors:	Allers, Thorsten Dattani, Ambika
Keywords:	DNA replication, Growth condition, Haloferax volcanii
Subjects:	Q Science > QH Natural history. Biology > QH573 Cytology
Faculties/Schools:	UK Campuses > Faculty of Medicine and Health Sciences > School of Life Sciences
Item ID:	65463
Depositing User:	McCulloch, Bryn
Date Deposited:	04 Aug 2021 04:42
Last Modified:	04 Aug 2021 04:42
URI:	https://eprints.nottingham.ac.uk/id/eprint/65463

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