Heichinger, C.; (2006) Characterisation of fission yeast DNA replication origins. Doctoral thesis , University of London.

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Abstract

In many eukaryotic organisms the chromosomal origins of DNA replication (ORIs) are not characterised by a clearly defined consensus sequence. In this thesis using the fission yeast, for the first time I have carried out a genome-wide analysis to identify such ORIs during the mitotic and meiotic cell cycles. The data can be summarised as follows: a total of 401 ORIs were identified which were used 29 percent of the time during mitotic S-phase and were spaced every 31 kilobases (kb) on average. The same ORIs were used during pre-meiotic S-phase although with lower efficiency in most chromosomal regions. A further 503 potential ORIs were used less efficiently at eight percent of the time during mitotic S-phase. This totals 904 ORIs which were distributed at an average inter-origin distance of 14 kilobases (kb) throughout 12.5 megabases (Mb) of the three chromosomes of fission yeast. These data support the idea of a continuum of ORI activity. The 401 efficient ORI loci contained A+T-rich regions located between genes, and these intergenic regions were typically larger than average. ORIs were not defined by a strict sequence consensus but the presence of AT-hook binding sequences. When the initiation factors Cdc18 and Cdt1 were over-expressed, regions of DNA containing particularly efficient ORIs with exceptionally large AT-hook binding domains became over-amplified, suggesting that interactions between these factors and efficient ORIs may be important for the mechanism ensuring that an ORI only fires once in each S-phase.

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