Errors in DNA replication can cause chromosome instability and gross chromosomal rearrangements (GCRs). For my thesis work I investigate how chromosome instability can originate in the telomere. Here I report how defects in Cdc13, a telomere specific protein, lead to chromosome instability and GCRs in Saccharomyces cerevisiae. Using a temperature sensitive mutant of Cdc13, I find that cdc13-induced instability can be induced in a single cell cycle and synergizes with replication stress (dNTP depletion via hydroxyurea). Additionally, I find that Cdc13 has to be functional during the cell’s S phase to suppress chromosome instability. Further genetic analysis suggests that that cdc13-induced chromosome instability depends on the generation of single stranded (ss)DNA, but not on the activity of canonical double strand break (DSB) repair pathways such as homologous recombination or non-homologous end joining. Finally, I demonstrate that telomeric unstable chromosomes can later progress and trigger rearrangements at centromeric loci. This system, using the conditional nature of the cdc13 mutation, promises a more complex analysis of the ontogeny of chromosome instability: in this case from errors semi-conservative DNA replication through the telomere to the formation and resolution of unstable chromosomes.

Herein we tested the repeatability of RAD-seq phylogeographic construction by creating a second, independent phylogeography of the pitcher-plant mosquito, Wyeomyia smithii. We sampled 25 populations drawn from different localities nearby previous collection sites and used these new data to construct a second, independent phylogeography to test the reproducibility of phylogenetic patterns. Our previous phylogeography was based on 3,741 phylogenetically informative markers from 21 populations and rooted with mitochondrial COI. The present phylogeography was based on 16,858 informative markers and rooted with RAD-seq. We found correspondence between clades at the extremes of W. smithii's distribution; however, there were several discrepancies between the trees, including the refugium that gave rise to all post-glacial populations. We observed that combining all 46 populations resolved these discrepancies and, equally importantly, that extensive taxon sampling in areas of historical importance is more valuable than increasing the number of informative sites in establishing an accurate, robust phylogeography. This thesis includes unpublished co-authored material.

The initiation of eukaryotic DNA replication occurs at replication origins, licensed by loaded double hexamers of Mcm2-7 proteins. The action of two kinases, DDK and S-CDK, triggers the loading onto Mcm2-7 of Cdc45 and the GINS complex to form the replicative CMG helicase. In yeast, the proteins Sld2 and Sld3 are CDK substrates that interact with Dpb11, and are required for the assembly of the CMG complex. Sld7 has been reported to associate with Sld3 throughout the cell cycle and regulate the function of Sld3 at replication initiation. Metazoan orthologues of these proteins have been identified: TopBP1, for Dpb11, is essential for replication;; Treslin, the Sld3 ortholog, is an essential S-CDK substrate that interacts with TopBP1 and is required for the CMG assembly;; in human cells MTBP has been found associated with Treslin, and is involved in the regulation of DNA replication, therefore MTBP as been proposed as a potential Sld7 ortholog. The aim of this project was to determine the role of MTBP in DNA replication using the Xenopus egg extract cell-free system. I found that MTBP and Treslin co-immunoprecipitate in metaphase and interphase extracts, suggesting that they form a complex throughout the cell cycle. MTBP, like Treslin, is recruited onto chromatin before the beginning of DNA replication and during S phase, independently of licensing and CDK/DDK activity. Immunodepletion of MTBP from extract leads to co- depletion of Treslin and causes a strong inhibition of DNA replication and impairment of CMG assembly. A partially purified fraction of extract, enriched for MTBP and Treslin, can rescue DNA replication in extracts depleted of MTBP. Analyses of the stoichiometry of the MTBP-Treslin complex has been carried out using gel filtration and sucrose gradient centrifugation; these identify a complex with an apparent molecular weight close to the one expected for a tetramer composed by two molecules of MTBP and two molecules of Treslin, resembling the model proposed for the interaction of the yeast proteins Sld3 and Sld7. These results, taken together, support the hypothesis that MTBP and Treslin form a multimeric complex that is required for the initiation of DNA replication and suggests that MTBP could be required to correctly position two Treslin molecules onto the MCM double hexamer to allow the bi-orientated firing of replication origins.

No description available.

Context. Software defect prediction is essential in reducing software development costs and in helping companies save their reputation. Defect prediction uses mathematical models to identify patterns associated with defects within code. Resources spent reviewing the entire code can be minimised by focusing on defective parts of the code. Recent findings suggest many published prediction models may not be reliable. Critical scientific methods for identifying reliable research are Replication and Reproduction. Replication can test the external validity of studies while Reproduction can test their internal validity. Aims. The aims of my dissertation are first to study the use and quality of replications and reproductions in defect prediction. Second, to identify factors that aid or hinder these scientific methods. Methods. My methodology is based on tracking the replication of 208 defect prediction studies identified in a highly cited Systematic Literature Review (SLR) [Hall et al. 2012]. I analyse how often each of these 208 studies has been replicated and determine the type of replication carried out. I use quality, citation counts, publication venue, impact factor, and data availability from all the 208 papers to see if any of these factors are associated with the frequency with which they are replicated. I further reproduce the original studies that have been replicated in order to check their internal validity. Finally, I identify factors that affect reproducibility. Results. Only 13 (6%) of the 208 studies are replicated, most of which fail a quality check. Of the 13 replicated original studies, 62% agree with their replications and 38% disagree. The main feature of a study associated with being replicated is that original papers appear in the Transactions of Software Engineering (TSE) journal. The number of citations an original paper had was also an indicator of the probability of being replicated. In addition, studies conducted using closed source data have more replications than those based on open source data. Of the 4 out of 5 papers I reproduced, their results differed with those of the original by more than 5%. Four factors are likely to have caused these failures: i) lack of a single version of the data initially used by the original; ii) the different dataset versions available have different properties that impact model performance; iii) unreported data preprocessing; and iv) inconsistent results from alternative versions of the same tools. Conclusions. Very few defect prediction studies are replicated. The lack of replication and failure of reproduction means that it remains unclear how reliable defect prediction is. Further investigation into this failure provides key aspects researchers need to consider when designing primary studies, performing replication and reproduction studies. Finally, I provide practical steps for improving the likelihood of replication and the chances of validating a study by reporting key factors.

This study used a quasi-experiment in order to evaluate the effect the SPOT-burglary profile on burglary arrest rates. A single police agency split into three different districts was used for the quasi-experiment. The SPOT-burglary profile was implemented in one district, while leaving the other two as control groups. The differences between the districts were controlled for using a statistical analysis. Burglary arrest rates were collected each month for all three districts for a period of one year before the implementation, and for six months after the implementation. Results show that the district who received the SPOT-burglary profile raised their burglary arrest rates by almost 75% in only 6 months, even after controlling for all relevant variables. This shows that the experimental intervention, the burglary profile, had a significant effect on the intended outcome- burglary arrest rates. The results of this study suggest that the SPOT-burglary profile may be able to provide law enforcement agencies with another tool to help increase burglary arrest rates in the future.

Thesis (Ph. D.)--University of Toledo, 2008. / Typescript. "Submitted as partial fulfillment of the requirements for the Doctor of Philosophy in Chemistry." Includes bibliographical references (leaves 245-252).

Thesis (Ph. D.)--Ohio State University, 2004. / Title from first page of PDF file. Document formatted into pages; contains xii, 208 p.; also includes graphics (some col.). Includes abstract and vita. Advisor: Deborah S. Parris, Dept. of Molecular Genetics. Includes bibliographical references (p. 191-208).

Thesis (Ph. D.)--University of Texas at Austin, 2003. / Supervisor: Makkuni Jayaram. Vita. Includes bibliographical references. Also available from UMI.

Drosophila embryos provide a rich source of replicative enzymes. Also, the duration of 2 hour embryo DNA synthesis phase of the cell cycle is approximately 6-fold shorter than the more regulated 9 hour embryo S phase. Thus, Drosophila embryos are a good system in which to explore the mechanisms and regulation of DNA replication. Early stage, 2 hour embryos contain at least two distinct DNA polymerases, DNA polymerase α and δ, as determined by associated enzymatic activities (DNA primase and 3'-5' exonuclease), inhibitor studies, immunologic reactivity, and processivity measurements. The observation that a δ-type enzyme with an inherent 3'-5' exonuclease activity is present in Drosophila embryos is a novel observation, and may have important implications for maintaining the fidelity of embryonic DNA synthesis. Both 2 hour and 9 hour embryos contained similar replicative activities. The enzymes which copurified with 2 hour and 9 hour DNA polymerases include a DNA primase activity with DNA polymerase α; and a 3'-5' exonuclease, 5'-3' exonuclease, and DNA ligase activities with DNA polymerase δ. The association of these activities suggests that DNA polymerase α-associated enzymes may initiate Okazaki fragments, which would then be elongated and ligated by the DNA polymerase δ-associated group.