Complex systems are those which exhibit one or more of the following inter-related behaviours: 1. Nonlinear behaviour: the component parts do not act in linear ways, that is the superposition of the actions of the parts is not the output of the system. 2. Emergent behaviour: the output of the system may be inexpressible in terms of the rules or equations of the component parts. 3. Self-organisation: order appears from the chaotic interactions of individuals and the rules they obey. 4. Layers of description: in which a rule may apply at some higher levels of description but not at lower layers. 5. Adaptation: in which the environment becomes encoded in the rules governing the structure and/or behaviour of the parts (in this case strictly agents) that undergo selection in which those that are by some measure better become more numerous than those that are not as “fit”. A single cell is a complex system: we cannot explain all of its behaviour as simply the sum of its parts. Similarly, DNA structures, social networks, cancers, the brain, and living beings are intricate complex systems. This thesis tackles all of these topics from a complex systems approach. I have skirted some of the philosophical issues of complex systems and mainly focussed on appropriate tools to analyse these systems, addressing important questions such as: • What is the best way to extract information from DNA? • How can we model and analyse mutations in DNA? • Can we determine the likely spread of both viruses and ideas in social networks? • How can we model the growth of cancer? • How can we model and analyse interactions between genes in such living systems as the fruit fly, cancers, and humans? • Can complex systems techniques give us some insight into the human brain? / http://proxy.library.adelaide.edu.au/login?url= http://library.adelaide.edu.au/cgi-bin/Pwebrecon.cgi?BBID=1290759 / Thesis (Ph.D.)-- School of Electrical and Electronic Engineering, 2007

Xanthomonas axonopodis pv citri (Xac), uma bactéria gram negativa, é a causadora da doença Cancro Cítrico que ocasiona enormes prejuizo na citricultura brasileira. Com o seqüenciamento do genoma deste patógeno foram encontradas inúmeras sequências codificadores com funções desconhecidas. Assim com a finalidade de estudos funcionais do genoma de Xac, foram gerados mutantes randômicos usando o transposon EZ::TN, que induziu disrupção aleatoria de seus genes com o intuito de avaliar que genes inativados afetam a patogenicidade da bactéria. Para o mapeamento do local de inserção do transposon foi desenvolvida uma metodologia baseada na técnica de PCR touchdown utizando oligonucleotídeos semidegenerados. A confiabilidade deste novo método foi comprovada através do mapeamento por Southem blot de alguns mutantes. Conseguiu-se mapear 90 mutantes randômicos com este método. Os testes de patogenicidade em citros mostraram mutantes com sua patogenicidade afetada observando-se variações nos sintomas da doença. Mutantes interessantes contendo uma ORF hipotética inativada ou tendo uma inserção num espaço intergênico foram achadas, sendo também detectados alguns mutantes com nocaute de genes nos seus plasmídeos pXac33 e pXac66. / Xanthomonas axonopodis pv citri (Xac), a gram-negative bacteria, is the causer of the Citrus Canker disease that produces enormous losses to the brazilian citrus sector. Many coding sequences with unknown functions were found in the genome sequence of this pathogen. Therefore, with the goal of functional studies of Xac\'s genome, random mutants using the EZ::TN transposon, have been generated, which carry aleatory disruptions of their genes, with the aim of evaluating inactived genes that potentially affect bacterial pathogenicity. A method based in the PCR touchdown technique using semidegenerate primers was developed for the mapping of the transposon insertion site. The reliability of this new method was tested by means of mapping some mutants using Southern blot. Ninety random mutants were mapped with this method. The pathogenicity tests in citrus showed mutants with their pathogenicity affected and variations in the disease symptoms were observed. Interesting mutants containing an inactive hypothetical ORF or with an insertion in intergenic regions have been found, and also some mutants with inactivated genes in their plasmids pXac33 and pXac66 were detected.

Xanthomonas axonopodis pv citri (Xac), uma bactéria gram negativa, é a causadora da doença Cancro Cítrico que ocasiona enormes prejuizo na citricultura brasileira. Com o seqüenciamento do genoma deste patógeno foram encontradas inúmeras sequências codificadores com funções desconhecidas. Assim com a finalidade de estudos funcionais do genoma de Xac, foram gerados mutantes randômicos usando o transposon EZ::TN, que induziu disrupção aleatoria de seus genes com o intuito de avaliar que genes inativados afetam a patogenicidade da bactéria. Para o mapeamento do local de inserção do transposon foi desenvolvida uma metodologia baseada na técnica de PCR touchdown utizando oligonucleotídeos semidegenerados. A confiabilidade deste novo método foi comprovada através do mapeamento por Southem blot de alguns mutantes. Conseguiu-se mapear 90 mutantes randômicos com este método. Os testes de patogenicidade em citros mostraram mutantes com sua patogenicidade afetada observando-se variações nos sintomas da doença. Mutantes interessantes contendo uma ORF hipotética inativada ou tendo uma inserção num espaço intergênico foram achadas, sendo também detectados alguns mutantes com nocaute de genes nos seus plasmídeos pXac33 e pXac66. / Xanthomonas axonopodis pv citri (Xac), a gram-negative bacteria, is the causer of the Citrus Canker disease that produces enormous losses to the brazilian citrus sector. Many coding sequences with unknown functions were found in the genome sequence of this pathogen. Therefore, with the goal of functional studies of Xac\'s genome, random mutants using the EZ::TN transposon, have been generated, which carry aleatory disruptions of their genes, with the aim of evaluating inactived genes that potentially affect bacterial pathogenicity. A method based in the PCR touchdown technique using semidegenerate primers was developed for the mapping of the transposon insertion site. The reliability of this new method was tested by means of mapping some mutants using Southern blot. Ninety random mutants were mapped with this method. The pathogenicity tests in citrus showed mutants with their pathogenicity affected and variations in the disease symptoms were observed. Interesting mutants containing an inactive hypothetical ORF or with an insertion in intergenic regions have been found, and also some mutants with inactivated genes in their plasmids pXac33 and pXac66 were detected.

This study aimed the study of ovarian follicular dynamics in a interovulatory period in nulliparous Santa Ines sheep carrying the polymorphism FecGE. Were used 21 sheep that previously genotyped for FecGE polymorphism by PCR-RFLP. The animals were divided into three groups (n = 7 / group) according to the genotype mutation: WW - non-mutant wild; EW . heterozygote and EE - homozygous. Follicular dynamics were monitored by ultrasound within a period interovulatory. The capture and measurement of the ovaries images were made every 24 hours until detection of estrus, being adjusted to every 12 hours until ovulation. It was found that 15 animals (71.42%) showed the emergence of three waves and six (28.58%) had four follicular waves. Don ft were influence of genotype on the selection, follicles . 2 and . 4 mm, (2,8 } 0,7) and follicular dominance, follicles > 5 mm, (0,9 } 0,1), the interovulatory period (17,5 } 0,2) and the duration of estrus (38,11 } 0,9). The largest number of ovulations was observed in homozygous sheep (2.4 } 0.2) compared to the wild type (1.7 } 0.2), however, heterozygous presenting intermediate value (1.9 } 0.1) but not differ from the others. Despite genetic groups did as not differ to the interval between the onset of estrus and ovulation (25,34 } 1,8), high individual variation was observed among sheep within each genotype. Santa Ines sheep that carriers the polymorphism FecGE express some differences in ovarian follicular dynamics in relation to non-mutants, being represented mainly by the largest number of ovulations in homozygous genetic group. / Este trabalho teve por objetivo o estudo da dinamica folicular ovariana do periodo interovulatorio em ovelhas nuliparas da raca Santa Ines portadoras do polimorfismo FecGE. Foram utilizadas 21 ovelhas previamente genotipadas para o polimorfismo FecGE por PCR-RFLP. Os animais foram divididos em tres grupos (n=7/grupo) de acordo com o genotipo relacionado a mutacao como segue: WW - selvagem nao mutante; EW - heterozigoto e; EE - homozigoto. A dinamica folicular foi acompanhada por ultrassom dentro de um periodo interovulatorio. A captura e mensuracao das imagens dos ovarios foram feitas a cada 24 horas ate a deteccao do estro, sendo ajustadas para cada 12 horas ate a ovulacao. Foi verificado que 15 animais (71,42%) apresentaram a emergencia de tres ondas e seis (28,58%) apresentaram quatro ondas de crescimento folicular. Nao houve influencia do genotipo sobre o numero de foliculos selecionados, . 2 e . 4 mm, (2,8 } 0,7) e dominantes, >5 mm, (0,9 } 0,1), o periodo interovulatorio (17,5 } 0,2) e, a duracao do estro (38,11 } 0,9). O maior numero de ovulacoes foi observado nas ovelhas homozigotas (2,4 } 0,2) em relacao as do tipo selvagem (1,7 } 0,2) sendo as heterozigotas apresentando valor intermediario (1,9 } 0,1), porem nao diferindo dos demais. Apesar dos grupos geneticos nao diferirem quanto ao intervalo entre o inicio do estro e a ovulacao (25,34 } 1,8 h), foi verificada alta variacao individual entre as ovelhas dentro de cada genotipo. Ovelhas nuliparas Santa Ines portadoras do polimorfismo FecGE expressam diferencas no tempo de sobrevivencia folicular e no diametro do foliculo ovulatorio em relacao as nao mutantes, sendo que o grupo genetico homozigoto apresenta um maior numero de ovulacoes.

Indiana University-Purdue University Indianapolis (IUPUI) / The LHX3 transcription factor is essential for pituitary gland and nervous system development in mammals. In humans, mutations in the LHX3 gene underlie combined pituitary hormone deficiency (CPHD) disease featuring deficits in anterior pituitary hormones and defects in the nervous system. The mechanisms that control temporal and spatial expression of the LHX3 gene are poorly understood. The proximal promoters of the human LHX3 gene are insufficient to guide expression in vivo and downstream elements including a conserved 7.9 kilobase (kb) enhancer region appear to play a role in tissue-specific expression in the pituitary and nervous system. In this study, I characterized the activity of this downstream enhancer region in regulating gene expression at the cellular level during development. Human LHX3 enhancer-driven Cre reporter transgenic mice were generated to facilitate studies of enhancer actions. The downstream LHX3 enhancer primarily guides gene transcription in αGSU-expressing cells secreting the TSHβ, LHβ or FSHβ hormones and expressing the GATA2 and SF1 transcription factors. In the developing nervous system, the enhancer serves as a targeting module for expression specifically in V2a interneurons. These results demonstrate that the downstream LHX3 enhancer is important in specific endocrine and neural cell types but also indicate that additional regulatory elements are likely involved in LHX3 gene expression in other cell types. Further, these studies demonstrate significant gonadotrope cell heterogeneity during pituitary development, providing insights into the cellular physiology of this key reproductive regulatory cell. The human LHX3 enhancer-driven Cre reporter transgenic mice provide a valuable tool for further developmental studies of cell determination and differentiation in the pituitary and nervous system. Furthermore understanding the regulation of human LHX3 gene will help develop tools to better diagnose and treat pituitary CPHD disease.