Thesis (Ph. D.)--University of Wisconsin--Madison, 1980. / Typescript. Vita. eContent provider-neutral record in process. Description based on print version record. Includes bibliographical references (leaves 115-124).

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"The Aurora Kinase proteins are a family of serine/threonine kinases that have been shown to play fundamental roles in controlling M phase progression in somatic cells. Aurora Kinase A protein is known to be vital for proper spindle assembly and therefore, chromosome segregation. Previous reports have shown that Aurora Kinase B is vital for proper completion of karyokinesis and cytokinesis in somatic cells. The role of Aurora Kinase C in somatic cells has been found to be less clear; however it appears to play an important role in spermatogenesis. Little is known about the role of these Aurora Kinase proteins mouse oocytes during oogenesis, and even less is known about them in embryos during early development. The objective of these studies was to characterize the presence, localization, and function of Aurora Kinase B and Aurora Kinase C protein and mRNA in mouse oocytes and early embryos. Oocytes and embryos were collected from hormone stimulated CF-1 mice and cultured for varying amounts of time. Cumulus denuded oocytes were either fixed for immunofluorescence microscopy studies, lysed for analysis of mRNA levels through the use of reverse transcription PCR (rtPCR) and quantitative rtPCR (q-rtPCR), lysed for protein analysis employing Western blotting, treated with Aurora Kinase protein inhibitor drugs, or microinjected with a siRNA pool targeting Aurora Kinase B. Samples were processed for immunofluorescence analysis using markers of spindle morphology (tubulins), Aurora Kinase B, Aurora Kinase C, and Aurora Kinase B activity (phospho Histone H3). Analysis of relative levels of Aurora Kinase B and Aurora Kinase C mRNA were assessed by rtPCR and q-rtPCR methods. Western blotting was performed on oocytes and early embryos to quantitate Aurora Kinase B and C protein levels. Aurora Kinase inhibitors, Hesperadin and ZM447439, were added to culture medium with mouse oocytes to determine the effects of the loss of Aurora Kinase activity. siRNAs were used to inhibit Aurora Kinase B mRNA in early embryos to ascertain the effect of functional loss of this transcript on embryo development. Marked differences were observed in the localization of Aurora Kinase B when unfertilized oocytes or pre-zygotic genome activation (ZGA) embryos were compared to post-ZGA samples. There was no evidence of Aurora Kinase B protein localized to the mitotic spindle or resultant midbody in oocytes and blastomeres of early embryos. Western blotting results supported this data. Embryos fixed post-ZGA demonstrated Aurora Kinase B localization at midbodies between dividing cells, as was found in mouse embryonic fibroblast control cells. Aurora Kinase C protein was not demonstrable in mouse oocytes, embryos, or control cells using immunocytochemistry or Western techniques. In contrast, Aurora Kinase B and Aurora Kinase C mRNAs were both found to be present in mouse oocytes and early embryos. q-rtPCR data further supported this finding for Aurora Kinase B and revealed that the mRNA level of this transcript is relatively constant until ZGA at which point a decrease relative to the earlier stages was observed. Transcript levels recovered post-ZGA and were comparable to the pre-ZGA levels. Functional inhibition of the Aurora Kinase family through the use of Hesperadin or ZM447439 demonstrated the importance of these proteins for proper microtubule and spindle organization, as these drugs disrupted both karyokinesis and cytokinesis in mouse oocytes and blastomeres of early embryos. Aurora Kinase B targeting siRNA also established a role for Aurora Kinase mRNA in embryos at the 2-cell stage based on the disruption of the cell cycle that was observed in treated embryos. Given earlier reports showing the vital role of the Aurora Kinase proteins in proliferating somatic cells, knowledge of the expression and localization of these proteins in oocytes and early embryos is vital for the understanding of cell cycle control during oogenesis and early embryogenesis. Our data indicate that Aurora Kinase B mRNA may also play a role in early embryogenesis, demonstrating a need for analysis of transcript as well as protein. Our results, as well as outcomes of future experiments suggested by our work, may provide significant insight into cell cycle regulation differences between somatic and embryonic cells. These differences may have a profound impact upon manipulated embryos including those reconstructed through somatic cell nuclear transfer. "

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Mitosis and meiosis are different forms of cell division. Mitosis is a non-reductive form of cell amplification whereby DNA chromosomes are replicated and segregated to form two progeny copies of the progenitor cell. Meiosis is a reductive form of cell division creating progeny containing half the chromosome copies of the progenitor cell. Improper chromosome segregation creates aneuploidy, which is poorly tolerated in cells. In cycling mitotic cells, aneuploidy leads to genome instability and cell death. Following meiosis, aneuploidy is associated with infertility, miscarriages, and birth defects. To segregate chromosome copies properly, pairs are physically organized and segregated to progeny cells by a mitotic spindle, whose functionality is tightly regulated. Kinesins are a family of highly conserved dimeric ATPase proteins which; organize spindle shape and size, facilitate chromosome capture and attachment to the spindle, and generate forces which are required for segregation. I investigated the molecular structure and function of human kinesin 13 family protein, Mitotic Centromere Associated Kinesin, MCAK. MCAK is a microtubule depolymerase whose full molecular structure and mechanism of depolymerization is not fully understood. Using in vitro biochemical assays and in vivo TIRF imaging, I found that altering MCAK molecular structure alters MCAK sub-spindle localization and by inference, alters global microtubule dynamics. This study suggests a potential mode for regulating of MCAK activity/function requiring further testing. Compared to over 30 kinesins in humans, showing a large amount of functional redundancy, yeast only has 6 identified kinesins whose function during meiotic cell division are still relatively unknown. I screened the importance and redundancy of yeast kinesins during meiosis. The results suggest similar roles and redundancies in meiosis to that during mitosis, despite different biochemical and biophysical spindle environments. Together, my investigations broaden the understanding of kinesin regulation and functional redundancy during different types of cell division.

Research Report submitted to the Faculty of Science, University of Witwatersrand, Johannesburg, in Partial Fulfilment of the Requirements for the Degree of Master of Science (Science Education). 2015. / In my Honours degree, I researched on learners’ understanding of meiosis after I had taught them the topic. The study was done on three schools. The results of the study revealed that the majority of learners from my school were unable to identify and explain some concepts in meiosis. The failure of the learners to understand the topic prompted me to reflect on my content knowledge of meiosis and its teaching. To investigate my own content knowledge in this self-study, I used concept maps and CoRes. Concept maps were seeking to develop my content knowledge and CoRes were seeking to develop both content knowledge and pedagogy of teaching meiosis. The aim of the self-study was therefore to improve my content knowledge and pedagogical content knowledge (PCK) of meiosis through the use of concept maps and CoRes as planning tools. The development of content knowledge and pedagogy was done with the help of collaborative friends. Of the two planning tools I used (concept maps and CoRes), three concept maps and three CoRes were constructed. Each of the concept maps was analysed using number of concepts identified and the number of propositions. Both qualitative and quantitative methods were used to analyse the concept maps. The number of both concepts and propositions gave an indication of the development of content knowledge from one concept map to the other. CoRes were analysed qualitatively using a framework. The framework used focused on curricular saliency, student prior knowledge, what makes the topic difficult or easy, teaching strategies and representations. From these aspects of the framework, the teachers’ content knowledge and pedagogy was identified to see whether there was development from one CoRe to the other. The study revealed that both concept maps and CoRes when used as planning tools can develop the teachers’ content knowledge and pedagogy on meiosis. Concept maps helps to indentify content gaps and misconceptions. CoRes helped me in the identification of the big ideas for the teaching of meiosis, the content which learners need to know and the identification of teaching strategies which can help the topic to be understood better. Recommendations from the study were that teachers should read about what they teach to improve content knowledge. Teachers should team up to produce teaching tools like CoRes.

Meiosis is a cell division process central to the life cycle of all sexual eukaryotic organisms. Chromosome pairing, genetic recombination and subsequent nuclear division during meiosis produces four genetically distinct haploid gametes from a single diploid cell. Allohexaploid wheat ( Triticum aestivum ) behaves meiotically as a diploid, despite the existence in the genome of three closely related ( homoeologous ) genomes, A, B and D. Chromosome pairing during prophase I of meiosis in wheat is restricted to true homologous chromosomes, the result being the formation of 21 bivalents at meiotic metaphase I. The genetic control of chromosome pairing in wheat is under the control of several pairing homoeologous ( Ph ) genes, located predominantly on chromosome groups 3 and 5. The major suppressors of homoeologous pairing are Ph1 and Ph2. Their cytogenetic effect has been intensively studied but at the molecular level little is known about their function. The isolation and characterisation of Ph genes from wheat would lead to greater understanding of chromosome pairing mechanisms in complex allopolyploids, and may enable development of effective strategies for alien gene introgression from related species to modern wheat cultivars. In this study, several genomics - based approaches were adopted to explore the expressed portion of the wheat genome in order to identify and characterise genes that could function in the molecular processes regulating meiosis. The first approach used comparative genetics to characterise the region deleted in the ph2a mutant ( a deletion mutant at Ph2 ). The rice genomic region syntenous to that deleted in the ph2a mutant was identified through comparative mapping and used in searches of wheat databases to identify ESTs with significant similarity. Southern analysis confirmed a syntenous relationship in the wheat and rice genomic regions and defined precisely the position of the breakpoint in ph2a. What seems to be a terminal deletion on 3DS is estimated to be approximately 80 Mb in length. We can tentatively predict the identification of approximately 220 genes from the region deleted in ph2a. The putative role of identified candidate Ph2 genes is discussed. The second approach explored the validity of recent proposals suggesting the presence of a meiotic gene cluster in the region of Ph2. The transcriptional characteristics of genes linked to Ph2 were investigated using data from wheat EST databases in combination with recently developed analysis software. The tissue - distribution of mRNAs derived from genes linked to Ph2 is shown to resemble that of other large chromosomal regions in the wheat genome. It is concluded that the apparently high number of genes from the Ph2 region expressed in wheat meiotic tissue is not indicative of a meiotic gene cluster in this region, but rather highlights the transcriptional complexity of meiotic anther tissue. Finally, the meiotic expression pattern of approximately 1800 wheat genes was examined using cDNA microarrays. Two approaches were taken. Firstly, the applicability of microarrays to identify differentially expressed genes between wild - type anthers and anthers of three Ph mutant genotypes was investigated. These experiments failed to reveal significant down - regulation of genes in Ph mutant anthers compared to wild - type. Possible explanations are discussed. Secondly, the expression of all microarray clones was examined from pre - meiotic interphase through to the tetrad stage of meiosis. A number of candidate wheat genes involved in meiotic and anther developmental processes have been identified and are discussed. Prior to this study, the methods available to identify wheat meiotic genes, in particular as candidates for Ph2, were limited. The recent development of genomics in plant biology provided an opportunity for a new approach towards gene discovery and genome structural analysis in relation to meiosis. This research illustrates the need for, and the effectiveness of a new approach to study meiosis, contributing to our knowledge of the structural and functional characteristics of genes linked to Ph2, and establishing a strong basis for further wheat meiotic gene characterisation. / Thesis (Ph.D.)--School of Agriculture and Wine, 2003.

Thesis (M.Sc.)--York University, 2008. Graduate Programme in Biology. / Typescript. Includes bibliographical references (leaves 54-66). Also available on the Internet. MODE OF ACCESS via web browser by entering the following URL: http://gateway.proquest.com/openurl?url_ver=Z39.88-2004&res_dat=xri:pqdiss&rft_val_fmt=info:ofi/fmt:kev:mtx:dissertation&rft_dat=xri:pqdiss:MR38792