L’autisme et la déficience mentale sont deux syndromes neuro-développementaux impliquant des facteurs génétiques. Notre travail a consisté à rechercher de nouveaux gènes candidats ou facteurs de susceptibilité chez 106 patients atteints d’autisme et 68 de déficience mentale non syndromique sporadique.Nous avons observé une association entre l’allèle 4 d’un marqueur microsatellite GXAlu localisé en 17q11.2 dans l’intron 27b du gène NF1 et des patients atteints de déficience mentale non-syndromique.Nous avons contribué à la mise en évidence d’une augmentation d’expression du transcrit NLGN4X, chez un patient autiste avec un retard mental non-syndromique présentant une mutation dans le promoteur du gène NLGN4X.L’étude de la région 22q13 par MLPA, nous a permis de mettre en évidence une délétion de novo d’au moins 1Mb chez un patient autiste.Les variations de nombre de copies (CNV) ont été étudiées chez des autistes par QPCR. Nous avons identifié 27 variations réparties sur 17 gènes parmi les 36 explorés. Les CNV observés dans les gènes ITGA6, TAGLN3, HOXA1, DLG4 et UBE2C sont intéressants en raison de l’implication de ces gènes dans le développement cérébral ou la fonction neuronale.L’ensemble de ces résultats nécessite des expériences complémentaires de validation. / Autism and mental retardation are two neurodevelopmental syndromes involving genetic factors. Our work consists in finding new candidate genes or susceptibility factors. 106 autistic patients and 68 sporadic non-syndromic mentally retardated patients were studied.We have shown an association between allele 4 of a microsatellite marker GXAlu locasized in 17q11.2, in intron 27b of the NF1 gene and patients with non-syndromic mental retardation.We contributed to the study on the NLGN4X gene. We demonstrated an increase of expression of NLGN4X transcript, in an autistic patient with non-syndromic mental retardation linked to a mutation in the NLGN4X gene promoter.We study the 22q13 region with MLPA method, we have demonstrated a deletion de novo of at least 1Mb in an autistic patient.The copy number variations (CNV) have been investigated in an autistic population by QPCR. We identified 27 variations on 17 genes among the 36 investigated. The CNV observed in ITGA6, TAGLN3, HOXA1, DLG4 and UBE2C genes are interesting because of the involvement of these genes in brain development or neuronal function.These results require further experiments for validation.

The nurse shark, Ginglymostoma cirratum, is a sedentary shark species that inhabits coral reefs in the tropical and subtropical Atlantic Ocean and along the western coast of the Americas in the Pacific Ocean. Nurse shark tissue samples were collected from the Bahamas, Belize, Brazil, and Dry Tortugas National Park in Florida. 186 individuals were genotyped at 11 microsatellite loci, the control region of the mitochondrial genome was sequenced in 190 individuals, and 89 individuals from the Bahamas, Belize, and Dry Tortugas were genotyped at the major histocompatibility complex (MHC) class IIα locus. An analysis of molecular variance (AMOVA) for the microsatellite loci indicated significant subdivision only between the Bahamas and Dry Tortugas populations. An AMOVA for the mitochondrial control region sequences indicated significant subdivision between all population pairs. The AMOVA for MHC class IIα locus indicated significant subdivision between two population pairs: the Bahamas population and the Dry Tortugas population and the Belize population and the Dry Tortugas population. The nurse shark has the lowest mitochondrial DNA nucleotide diversity (π=0.0125%) and haplotype diversity (h=0.2402) of any shark species to date. There were 14 MHC alleles from 39 polymorphic sites; ten were the same as published alleles (Kasahara et al. 1993; Ohta et al. 2000). This study was the first study to use MHC class IIα genes as a marker for population genetics in sharks. Our results showed that MHC class IIα locus behaves as a diploid locus and is a powerful tool for determining population genetic structure between populations.

Made available in DSpace on 2014-06-11T19:26:08Z (GMT). No. of bitstreams: 0 Previous issue date: 2008-07-25Bitstream added on 2014-06-13T20:54:10Z : No. of bitstreams: 1 demore_ps_me_jabo.pdf: 1002271 bytes, checksum: a27ac944079c717a8ba5366fe7e35456 (MD5) / Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq) / Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES) / O oídio em soja, trata-se de uma doença praticamente presente em todos os paises produtores. Os marcadores moleculares microssatélites ou SSR (Simple Sequence Repeat) têm sido amplamente utilizados no processo de seleção assistida de genótipos de soja. Assim, o objetivo deste estudo foi obter marcadores microssatélites próximos ao gene de resistência ao oídio em soja. O estudo foi realizado em duas populações F2, oriundas de cruzamentos entre parentais contrastantes quanto à resistência ao oídio. Para o estudo, foram selecionados marcadores microssatélites a uma distância de até 42 cM ao redor do gene Rmd (resistência ao oídio). Utilizou-se o método de BSA (Bulked Segregant Analysis) na avaliação dos marcadores, para a comparação com a análise fenotípica das populações. Na análise foram utilizados dez iniciadores SSRs para as duas populações, sendo identificados quatro marcadores polimórficos para o cruzamento 1 (MGBR95-20937 x IAC-Foscarin 31) e três para o cruzamento 2 (MGBR 46/Conquista x EMBRAPA 48). Pela análise de Qui-quadrado da avaliação fenotípica, confirmou-se à segregação esperada (3:1) de um gene dominante condicionando a resistência. Os marcadores polimórficos também segregaram conforme o esperado (1:2:1) já que possuem natureza codominante. Para as populações 1 e 2, os melhores resultados foram obtidos com os marcadores Sat_366 e Sat_393, respectivamente, localizando-se a 9,41 e 12,45 cM de distância do gene, sendo considerados promissores na seleção assistida para resistência ao oidio em soja. / Powdery mildew in soybeans, it is a disease present in virtually all producing countries. The molecular markers microsatellites or SSR (Simple Sequence Repeat) have been widely used in the assisted selection of soybean genotypes. The objective of this study was to obtain microsatellites markers near the gene for resistance to powdery mildew in soybeans. The study was conducted in two populations F2, from crosses between contrasting parents about the resistance to powdery mildew. For the study, were selected microsatellites markers at a distance of 42 cM around the gene Rmd (resistance to powdery mildew). It was used the method of BSA (Bulked Segregant Analysis) in the evaluation of markers, for comparison with the phenotypic analysis of populations. In the analysis were used in ten initiators SSRs for the two populations, and identified four polymorphic markers for the crossing 1 (MGBR95-20937 x-IAC Foscarin 31) and three for the crossing 2 (EMBRAPA MGBR 46/Conquista x 48). For the analysis of chi-square of the phenotypic evaluation, it is confirmed the segregation expected (3:1) of a dominant gene conditioning the resistance. The polymorphic markers also segregation as expected (1:2:1) that have already codominante nature. For the populations 1 and 2, the best results were obtained with the markers Sat_366 and Sat_393, respectively, finding itself to 9.41 and 12.45 cM distance of the gene and are considered promising in assisted selection for resistance to soybean in powdery mildew.

Made available in DSpace on 2015-03-26T13:42:12Z (GMT). No. of bitstreams: 1 texto completo.pdf: 831305 bytes, checksum: 7de0dccf59e4128a858ec0de24b05804 (MD5) Previous issue date: 2009-08-03 / Coordenação de Aperfeiçoamento de Pessoal de Nível Superior / Common bean (Phaseolus vulgaris L.) is a crop of great nutritional, economical and social importance. Breeding programs use molecular markers as important auxiliary tools for various types of genetic studies. Different classes of molecular markers have been developed, and among them microsatellites highlight. Microsatellites are DNA simple sequence repeats (SSR) distributed in tandem along the genome, forming highly variable polymorphic sites, enabling their use as molecular markers. SSRs are codominant, multiallelic, and thus can be used in several types of studies, mainly for genetic mapping. Primers flanking microsatellite sequences are commonly developed from genomic libraries, enriched genomic libraries, sequences obtained from databases and, alternatively, from internal simple sequence repeats (ISSR-PCR). Common bean breeding molecular geneticists have developed SSR markers for mapping specific traits of interest. However, a saturated consensus genetic map for common bean has not been established so far that can be used as a reference for the development of specific maps. Therefore, the BIOAGRO/UFV common bean breeding program developed a population of recombinant inbred lines (RILs) which is suggested to be used to integrate, in one single map, all microsatellite markers that have been developed so far. However, to saturate the map, there must be a great number of markers available. The objective of the present study was to develop and validate primers that amplify microsatellites sequences obtained from enriched genomic libraries and from ISSR sequences. In the first case, two enrichedgenomic libraries for microsatellite sequences, that had been developed in a previous study, were used. In the present study, 207 clones were selected from these two genomic libraries. One hundred and ninety six of these clones (94.68%) were sequenced and 184 (88.89%) of them could be analyzed, 133 clones from library 1 and 51 from library 2. Forty eight redundant clones (26.09%) were detected. Clone analysis led to the identification of 66 (49.62%) microsatellite motifs in library 1 and 20 (39.22%) in library 2, and 56 primer pairs were designed. From the 56 primer pairs developed, 34 were characterized andtested in 20 Mesoamerican and Andean genotypes, including AND277 and Rudá. All the primer pairs were able to generate PCR products and six (17.65%) generated polymorphic DNA bands among the tested genotypes. In the ISSR enrichment methodology, 250 clones were seleted with sizes over 400 bp. From these 250 clones, 168 (67.2%) were sequenced and 103 (41.2%) could be analyzed. Thirty redundant clones (29.13%) were detected. Clone analyses led to the identification of 58 microsatellite motifs (56.31%) and 32 primer pairs were developed. Out of these, 10 were characterized and tested in the same genotypes used in the previous methodology. Out of the 10 primer pairs tested, six were identified as codominant markers and the other four as dominant. The codominant markers revealed no polymorphisms among the tested genotypes. Additionally, microsatellite containing sequences obtained from both methodologies were submitted to BLAST analysis against sequences deposited in public databases. Similarity was identified between the SSR sequences and transcribed and non-transcribed regions, from nuclear, mitochondrial and chloroplast genomes, and also with retrotransposon sequences. Both methodologies were effective for selecting, in common bean, sequences that contain microsatellites. The results obtained represent an initial effort to select molecular markers that will be mapped in the future RIL's consensus population, contributing for the construction of a satured genetic map for the species. In addition, these primers can be used in different types of genetic studies which are important for common bean breeding programs that use molecular markers. / O feijão-comum (Phaseolus vulgaris L.) é uma cultura de destacada importância nutricional, econômica e social. Programas de melhoramento genético do feijoeiro têm utilizado marcadores moleculares como importantes ferramentas auxiliares em diversos tipos de estudos genéticos. Diferentes classes de marcadores têm sido desenvolvidas, dentre as quais se destacam os microssatélites. Os microssatélites (SSR) são seqüências simples repetidas de DNA, que se repetem em tandem ao longo do genoma, formando sítios altamente polimórficos, o que possibilita o seu uso como marcas moleculares. Como marcadores, são codominantes, multialélicos, e aplicáveis em diversos tipos de estudos, principalmente no mapeamento genético. Primers que flanqueiam sequências SSR geralmente são desenhados a partir da construção de bibliotecas genômicas, bibliotecas genômicas enriquecidas, sequências depositadas em bancos de dados e, alternativamente, a partir de seqüências internas simples repetidas (ISSR). Geneticistas moleculares têm desenvolvido marcadores SSR com o intuito de mapear genes que codificam determinadas características de interesse. Entretanto, não existe um mapa consenso saturado para o feijão que sirva como referência para auxiliar na construção de mapas específicos. Nesta perspectiva, o Programa de Melhoramento Genético do Feijoeiro do BIOAGRO/UFV desenvolveu uma população de RIL s que poderá ser usada para integrar, em um único mapa, todos os marcadores SSR já desenvolvidos. No entanto, para a saturação do mapa, há necessidade de um grande número de marcadores. O presente trabalho teve o objetivo de desenvolver e validar primers que amplifiquem regiões contendo microssatélites a partir da metodologia da construção de bibliotecas genômicas enriquecidas e a partir de ISSR. Na primeira metodologia, em trabalho anterior, foram construídas duas bibliotecas genômicas enriquecidas para seqüências SSR. No presente trabalho, a partir das bibliotecas genômicas desenvolvidas, foram selecionados 207 clones contendo insertos de tamanho desejado. Destes, foram seqüenciados 196 (94,68%), dos quais 184 (88,89%) puderam ser analisados, sendo 133 clones da biblioteca 1 e 51 da biblioteca 2. Foram detectados 48 (26,09%) clones redundantes. A análise dos clones permitiu identificar 66 (49,62%) motivos SSR na biblioteca 1 e 20 (39,22%) na biblioteca 2, a partir dos quais foram desenhados 56 pares de primers. Destes, 34 tiveram suas condições de amplificação otimizadas e padronizadas e foram testados quanto ao polismorfismo detectado entre 20 genótipos andinos e mesoamericanos, incluindo os genitores AND277 e Rudá. Todos os primers geraram produtos de amplificação e seis (17,65%) amplificaram produtos polimórficos entre os genótipos testados. Em relação à metodologia de enriquecimento por ISSR-PCR foram selecionados 250 clones contendo insertos com tamanho desejado, obtidos a partir da amplificação por ISSRPCR, clonagem dos fragmentos e transformação de células competentes. Dos 250 clones, 168 (67,2%) foram sequenciados e 103 (41,2%) puderam ser analisados. Foram detectados 30 clones redundantes (29,13%). A análise das sequências permitiu identificar 58 motivos microssatélites (56,31%) e foi possível o desenho de 32 pares de primers. Destes, 10 tiveram suas condições de amplificação padronizadas e foram analisados quanto ao polimorfismo detectado entre os mesmos 20 genótipos andinos e mesoamericanos utilizados na metodologia de bibliotecas genômicas enriquecidas. Dos 10 pares de primers testados, seis comportaram-se como marcadores codominantes e quatro como dominantes. Dos codominantes nenhum mostrou-se polimórfico dentre os genótipos testados. Adicionalmente, as sequências contendo motivos microssatélites, obtidas a partir das duas metodologias utilizadas, foram submetidas à busca por similaridade com sequências já caracterizadas em bancos públicos de sequências. Foi identificada similaridade com regiões transcritas e não traduzidas, e com regiões codificadoras de proteínas, a partir do genoma nuclear, mitocondrial e do cloroplasto, e também a partir de sequências advindas de retrotransposons. As duas metodologias utilizadas foram eficazes para a seleção, no feijoeiro, de seqüências contendo microssatélites. Estes resultados representam um primeiro esforço no sentido de selecionar marcadores moleculares que serão futuramente mapeados na população consenso de RILs, além de fornecer marcadores que poderão ser usados nos mais variados tipos de estudos genéticos, contribuindo de fundamental maneira para o aprimoramento dos programas de melhoramento do feijoeiro comum que utilizam marcadores moleculares.

Made available in DSpace on 2015-04-20T12:31:34Z (GMT). No. of bitstreams: 1 Cleiton Fantin.pdf: 1078270 bytes, checksum: 5ca25ef1341a3ab0c4b469d3442db3c8 (MD5) Previous issue date: 2008-01-07 / Coordenação de Aperfeiçoamento de Pessoal de Nível Superior / Few data are available about reproductive biology of the genus Podocnemis, especially with respect to the type of mating system. To date studies of mating system with species of the genus Podocnemis have been done only with P. expansa. In the present study we developed 18 primer pairs specific to microsatellite regions in P. unifilis, and we tested their transferability to other species of the genus Podocnemis (P. expansa, P. sextuberculata, P. erythrocephala, P. vogli and P. lewyana) and also to Peltocephalus dumerilianus. These microsatellite markers have proven to be powerful molecular tools for investigating mating systems of P. unifilis, P. sextuberculata and P. erythrocephala. Using different microsatellite panels a minimum of two contributing parents were found in nests of P. erythrocephala, and the presence of up to three contributing parents was found in nests of P. unifilis and P. sextuberculata. These results directly contribute to the knowledge of the reproductive system and indirectly to studies related to the impact of management and reproduction in nature as well as in captivity. Moreover, our results will contribute to improvements in captive breeding programs. / Poucos dados estão disponíveis com relação à biologia da reprodução no gênero Podocnemis, mais especificamente com relação ao sistema de reprodução. Até hoje os estudos sobre sistema de reprodução realizados no gênero Podocnemis foram realizados somente com P. expansa. No presente trabalho desenvolvemos 18 iniciadores específicos para regiões microssatélites em P. unifilis, e testamos sua transferibilidade nas outras espécies do gênero Podocnemis (P. expansa, P. sextuberculata, P. erythrocephala, P. vogli e P. lewyana) e também em Peltocephalus dumerilianus. Estes microssatélites apresentaram um grande potencial como marcadores moleculares para investigar o sistema de reprodução em P. unifilis, P. sextuberculata e P. erythrocephala. Utilizando diferentes grupos de locos de microssatélites, encontrou-se o mínimo de dois pais contribuindo para as ninhadas de P. unifilis, e o mínimo de três pais contribuindo para as ninhadas de P. sextuberculata e P. erythrocephala. Esses resultados contribuem diretamente para o conhecimento do sistema de reprodução e indiretamente para estudos relacionados ao impacto de manejo e reprodução tanto na natureza como em cativeiro destas espécies. Além disso, nossos resultados poderão contribuir para um melhoramento nos programas de reprodução em criadouros.

O avanço na distribuição geográfica de mosquitos vetores é seguido pela emergência de vírus e doenças em novas áreas para as quais não há disponibilidade de vacinas efetivas e drogas terapêuticas específicas são insuficientes. Métodos de controle de mosquitos tradicionais perderam efetividade, devido principalmente a grande capacidade reprodutiva e flexibilidade genômica dos mosquitos. Controle químico cada vez mais tornase restrito, acarretando na urgente necessidade de novas formas de controle. A liberação de machos carregando um gene letal dominante (RIDL) oferece novas abordagens aplicáveis ao controle de mosquitos e ainda assim ecológicas e espécie específica. Mosquitos Culex quinquefasciatus foram transformados com sucesso apenas uma vez, apesar do esforço de diversos laboratórios em obter uma linhagem transgênica estável. Foi desenvolvido um método de expressão transiente em mosquitos Culex, que insere plasmídeos contendo genes efetores na hemolinfa e tecidos subjacentes do mosquito. Foi observada a expressão da proteína fluorescente DsRed2, em mosquitos Culex quinquefasciatus adultos mediada por plasmídeos. Esta expressão pode ser considerada um importante passo na transformação de mosquitos Culex, além de potencial uso em estratégias de controle genético e interações gênicas. Para que novas formas de controle sejam realmente efetivas é vital que se conheça a estrutura genética da população alvo. Marcadores moleculares têm sido extensivamente utilizados em estudos filogenéticos e taxonômicos de diversas espécies de insetos. Microssatélites são de grande utilidade para observar estruturas populacionais, tanto em âmbito geográfico, quanto na escala evolucionária. Foi possível observar a formação de clusters e de padrões genéticos distintos entre as populações analisadas, criando um panorama genético dos mosquitos Culex quinquefasciatus coletados no Brasil / The increase in the geographic distribution of vectors is accompanied by the emergence of viruses and diseases in new areas. There are insufficient specific therapeutic drugs available and there are no reliable vaccines to malaria or dengue. Most mosquito control measures have failed to achieve their goals, mostly because of the mosquitos great reproductive capacity and genomic flexibility. Chemical control is increasingly restricted therefore other strategies for mosquito control are desperately needed. Releasing of Insects Carrying a Dominant Lethal Gene (RIDL) offers a new approach that can be applied to mosquitoes yet environmentally friendly and species-specific. Culex quinquefasciatus mosquitoes have been successfully genetically modified only once, despite the efforts of several laboratories to transform and establish a stable strain. We have adapted a transient gene expression method, in Culex, that delivers plasmid DNA directly to the mosquito hemolymph and additional tissues. We were able to express DsRed2 fluorescent protein in adult Culex quinquefasciatus mosquitoes by injecting plasmids directly into their thorax. The expression of DsRed2 in adult Culex quinquefasciatus mosquitoes is an important stepping stone to genetic transformation and the potential use of new control strategies and genetic interactions. If new methods of control are to become really effective it is vital to know the genetic structure of the target population. Molecular markers have been widely used in phylogenetic and taxonomic studies of various groups of insects. Microsatellites are very useful for observing the population structure both geographically and evolutionarily. It has been possible to observe the formation of clusters and distinct genetic patterns among the analyzed populations creating a panorama of Culex quinquefasciatus mosquitoes populations present in Brazil

Submitted by Renata Lopes (renatasil82@gmail.com) on 2016-09-15T13:54:35Z No. of bitstreams: 1 karlagasparinidossantos.pdf: 1186792 bytes, checksum: dac7f21d89dd87a474e971fead9a7387 (MD5) / Approved for entry into archive by Diamantino Mayra (mayra.diamantino@ufjf.edu.br) on 2016-09-26T20:20:26Z (GMT) No. of bitstreams: 1 karlagasparinidossantos.pdf: 1186792 bytes, checksum: dac7f21d89dd87a474e971fead9a7387 (MD5) / Made available in DSpace on 2016-09-26T20:20:26Z (GMT). No. of bitstreams: 1 karlagasparinidossantos.pdf: 1186792 bytes, checksum: dac7f21d89dd87a474e971fead9a7387 (MD5) Previous issue date: 2011-08 / Em países de clima tropical, os prejuízos causados pelo carrapato Riphichephalus (Boophilus) microplus causam grande impacto nos sistemas de produção bovino. A identificação de regiões genômicas e marcadores de DNA associados à resistência ao parasita poderão ser utilizados como estratégia para seleção de animais mais resistentes. Já foram descritos sete QTL relacionados à resistência ao carrapato bovino, porém, os estudos iniciais geralmente detectam QTL com uma resolução baixa ou moderada, sendo necessário o posterior refinamento da região onde o mesmo foi detectado. Dessa forma, esse trabalho teve como objetivo refinar a posição dos QTL previamente identificados com a adição de marcadores microssatélites em uma população bovina F2 formada a partir do cruzamento entre animais Gir e Holandês. Amostras de sêmen e sangue foram submetidas à extração de DNA e posterior PCR com os marcadores microssatélites. Os produtos de PCR foram detectados utilizando o seqüenciador automático de DNA MegaBACE 1000 e as análises estatísticas foram feitas utilizando o software GridQTL. As análises realizadas nos cromossomos 2, 5, 10, 11, 21, 23 e 27 confirmaram a presença dos QTL encontrados anteriormente e reduziram significativamente os intervalos de confiança da maioria dos QTL. Os QTL encontrados no BTA 2 e 27 foram significativos a Pc<0.05, aqueles localizados no BTA 5, 10, 11, 21 e 23 foram significativos a Pc<0.01. O BTA 2 sofreu redução no intervalo de confiança de 12 cM passando de 22 para 10 cM, assim como o BTA 5 que passou de 20 para 8 cM. No BTA 10 foi confirmada a presença de dois picos de QTL com intervalo de confiança de 27cM anteriormente detectado em 47 cM. O BTA 11 passou a apresentar um intervalo de confiança de 19 cM e BTA 21 de 36 cM sofreu redução para 26 cM. O BTA 23 apresentou a menor redução no intervalo de confiança na estação de chuva passando de 17 para 14 cM e na seca manteve inalterado com I.C. de 12 cM, assim como o BTA 27 que sofreu redução de apenas 1 cM passando para 7 cM. A adição de marcadores e a redução no intervalo de confiança de QTL previamente encontrados é um importante passo para a identificação de genes relacionados à resistência ao carrapato. / In tropical countries, losses caused by tick Rhipichephalus (Boophilus) microplus causes a great impact on cattle production systems. The identification of genomic regions and DNA markers associated to the parasite resistance may be used to select resistant animals. Seven QTL associated with tick resistance were described, however, the initial studies detect QTL with low or moderate resolution, being necessary the refinement of the regions where the QTL were detected. Thus, the aim of this work was refine the position of QTL previous identified with addiction of microsatellite markers in Gir X Holstein F2 population. Semen and blood samples were submitted to DNA extraction and then, the PCR were done using microsatellite markers. The PCR products were detected using DNA automatic sequencer MegaBACE1000 and the statistical analysis was performed using GridQTL software. The analysis performed on chromosome 2, 5, 10, 11, 21, 23 and 27 confirmed the presence of QTL previously found and the confidence intervals were significantly reduced in most of QTL. The QTL found on the BTA 2 and 27 were significant at Pc<0.05 and those located on BTA 5, 10, 11, 21 and 23 were significant at Pc<0.01. The BTA 2 was reduced 12 cM in the C.I., from 22 to 10 cM, and BTA 5 from 20 to 8. In BTA 10 was confirmed the presence of two QTL peaks with C.I. 27 cM, previously detected in 47 cM. BTA 11 reduced C.I. to 19 cM and BTA 21 reduced from 36 cM to 26 cM. BTA 23 showed the smallest reduction in C.I. in the rainy season, from 17 to 14 cM and in dry season the C.I remained unchanged with 12 cM, as the BTA 27 that reduced only 1 cM reaching 7 cM. The addiction of markers and the reduction in the confidence interval of QTL previously found is an important step to identify genes related to ticks resistance.

Abstract The pattern and the genetic basis of variation in courtship song of D. virilis were studied using three different approaches: a candidate gene, a biometrical and a quantitative trait locus (QTL) method. Nucleotide variation in a candidate song gene, no-on-transientA, was analysed both within the species (D. virilis and D. littoralis) and between the species of the D. virilis group. Nucleotide variation showed no signs of selection and there was no association between the nucleotide or repeat length variation in nonA gene region and the song characters of the D. virilis group species. Molecular markers (microsatellites) were isolated for D. virilis and their cross-species amplification was tested in all members of the D. virilis group. Intraspecific variation in D. virilis was studied at the phenotypic level in male song characters and at the genetic level in microsatellites. Significant geographic variation was detected in both levels, grouping the strains according to the main continents of the species' distribution range: America, Asia, Europe and Japan. The strains with most extreme song phenotypes were chosen for further analysis. The inheritance of two courtship song characters, the number of pulses in a pulse train (PN) and the length of a pulse train (PTL) was studied by analysing the means and variances of these characters between parental and reciprocal F1, F2 and backcross males. This biometrical analysis showed the genetic basis of these song characters to be polygenic with significant dominance, epistatic and Y-chromosomal effects on both characters. A subset of these data (F2 generation males) were used to conduct a QTL study with the aid of a recombination linkage map constructed for the microsatellites. Composite interval mapping (CIM) revealed significant QTLs, which were shared in both characters. Altogether, significant QTLs, located on the X, 2nd, 3rd and 4th chromosome, were found to affect PN, whereas only QTLs on the 3rd chromsome was found to affect PTL. The effect of the same QTL on the 3rd chromosome on both characters accounted for 31.8% and 49.1% of the mean difference between the parental strains in PN and PTL, respectively. These results suggest the genetic basis for these song characters is caused mainly by autosomal QTLs with a relatively large effect.

Masters of Science / Genetic analysis for resistance to Woolly Apple Aphid in apple rootstock breeding populations MC Selala MSc Thesis, Department of Biotechnology, Faculty of Science, University of the WesternCape. The Woolly Apple Aphid (WAA) Eriosoma lanigerum (Hausm.) (Homoptera: Aphididae) is economically one of the most important pests in apple commercial production in the Western Cape province, South Africa. The apple cultivar Northern Spy possesses a single major gene (Er1) responsible for E. lanigerum resistance. This cultivar has been used as a commercial rootstock in apple breeding programmes. There are other genes also implicated in resistance to E. lanigerum from other cultivars. Manipulation and pyramiding of the E. lanigerum resistance genes (Er1, Er2 and Er3) might provide a necessary control for commercial apple production. The aim of this study was to construct a genetic linkage map for apple using microsatellite markers. The use of marker-assisted selection would greatly benefit local apple breeding programmes. Ninety six seedlings from a Northern Spy × Cox Orange Pippin mapping population were used for genetic linkage construction. Phenotypic data collection and analysis were performed to determine the E. lanigerum infestation patterns and the levels of resistance conferred by the Er1 gene from Northern Spy using 52 in vitro propagated seedlings in the greenhouse. Classification and quantification analysis showed association patterns between first assessments (30 days) to second assessment (60 days) in all replicate blocks. Roots and shoots data showed that it could be useful in quantitative trait loci (QTL) analysis, but may be used in different QTLs beingidentified due to the variations between roots and shoots data. A preliminary linkage map was constructed using a mapping population from Northern Spy × Cox Orange Pippin (96 seedlings).Fluorescently labelled published and predicted microsatellite markers were used in map construction. Primers were optimised using single apple cultivar and the detection of polymorphisms using nine apple cultivars. Optimised markers were multiplexed for high throughput data generation using the Polymerase Chain Reaction (PCR) technique. Multiplexed PCR products were pooled and analysed on an ABI 310 PRISM™ Genetic Analyser to determine allele fragment sizes, and the inherited segregation types in the seedlings. Computer software GenoTyper® 2.5.2 and JoinMap® 3.0 was used in data analysis from ABI 310 PRISM™Genetic Analyser and linkage map construction. Seventy two markers were used in linkage map construction, which produced nine linkage groups with some segments from the same linkage group. Twenty-one markers were aligned on the map 20 published and one predicted. Only one linkage group consisted of five markers while other linkage groups had two markers each. This study has proved that th preliminary linkage map could be used as the basis of a complete linkage map of Northern Spy × Cox Orange Pippin.

Methods to determine genetic diversity and relatedness within populations are essential tools for proper wildlife management. Today the approach of choice is polymerase chain reaction-based microsatellite analysis. Seven new polymorphic loci were isolated from a microsatellite-enriched Caribbean flamingo genomic library and used to characterize survey populations of Caribbean and African greater flamingos. In addition, four of these loci were used to verify parentage relationships within a captive-breeding population of African greater flamingos. Parentage predictions based upon gamekeeper observations of breeding and nesting did not always agree with genetic-based parentage analyses of the nine suggested family groups. Four family groups were supported (groups I, II, III and VI) by there results. However, an analysis of the remaining five suggested groups, with a total of eight offspring/dam and eight offspring/sire suggested relationships, yielded seven exclusions of the suggested dam and six exclusions of the suggested sire. This put the overall suggested dam exclusion rate at 35% and exclusion rate for suggested sires at 29%. Although the keeper observation data for our family groups must be considered a variable of concern at this time, these findings are certainly suggestive that more carefully controlled studies may reveal that flamingos are not monogamous as long accepted, but rather socially monogamous or even promiscuous. Thus we have now been able to both characterize and demonstrate the utility of our polymorphic microsatellite loci. We hope these results will interest additional wildlife facilities in further parentage and behavioral studies that will collectively aid to improve monitoring and maintenance of genetic diversity, and as provide better insight into breeding habits of both wild and captive populations.