No description available.

0009014A Msc thesis Science Animal plant and environmental sciences / Anopheles arabiensis is considered an important vector of human malaria in the southern African region where the disease is the major cause of morbidity and mortality. Gene flow plays an important role in malaria control with the spread of insecticide resistance. The main objectives of this study were to (i) measure the genetic variability within and between five populations (Botswana, Namibia, South Africa, Swaziland and Zimbabwe) of wild An. arabiensis and (ii) estimate the level of gene flow between natural populations across the Southern-limits of An. arabiensis. A total of 1225 An. arabiensis specimens were identified out of 1300 mosquitoes collected from 2000-2003 with the sample sizes ranging from 180-292 per country. Variation at four microsatellite markers was investigated on non-denaturing polyacrylamide gels. The results showed fewer variations between populations (2.96%) than within populations (82.60%) suggesting considerable homogeneity. However, estimates of gene flow (Nm) calculated from mean FST and RST statistics were relatively low, 1.14 and 1.19 respectively, suggesting somewhat restricted gene flow between populations. The occurrence of gene flow within subpopulations of An. arabiensis in Zimbabwe but not in South Africa is interesting with regard to the spread of insecticide resistance in Zimbabwe. The results presented here are obviously subject to the limitations inherent in manual, silver staining method of analysing microsatellite DNA markers. It is possible that a different set of results would be obtained if an Automated Sequencing Analyzer were used. ii

Fungi are present everywhere in indoor and outdoor environments. Many fungi are toxigenic or pathogenic that may cause various public health concerns. Rapid detection, quantification and characterization of fungi in living and working environments are essential for exposure risk assessment to safe guard public health. Rapid and accurate detection and identification of fungi using molecular method require specific markers. In this thesis, partial mt SSU and LSU rDNA were amplified and sequenced from 31 fungal species of 16 genera. Sequence alignments showed that fungal mt SSU and LSU rDNA contained sufficient amount of variation for the development of markers that can discriminate even among closely related species. Forty-eight probes were designed and were verified as highly specific to 25 fungal species commonly detected in living and working environments. These specific probes would have potential applications in clinical diagnosis and public health-related environmental monitoring. Nested PCR is a highly sensitive and specific method. Based on the nuclear 18S rDNA sequence variation pattern, three nested PCR systems were developed to detect the conifer tree pathogen Gremmeniella abietina, an ascomycete fungus that causes stem canker and shoot dieback in many conifer species. The three nested PCR systems showed high specificity and sensitivity. These methods could have broad applications in forest protection and disease management programs. Quantitative real-time PCR offers the ability of simultaneous detection and quantification of DNA of a specific microbe in one reaction. Based on the 18S rDNA sequence, two real-time PCR assays were developed to detect and quantify Wallemia sebi, a deuteromycete fungus commonly found in agricultural environments and is suspected to be a causative agent of farmer’s lung disease. Both PCR systems proved to be highly specific and sensitive for W. sebi detection even in a high background of other fungal DNAs. Application of the real-time PCR methods in the quantification of W. sebi in the aerosols of a farm revealed a high concentration of W. sebi spores (107/m3). The study indicates that W. sebi is a dominant fungus in agriculture environments. Cladosporium spores are important aeroallergens, and prolonged exposure to elevated spore concentrations can provoke chronic allergy and asthma. A TaqMan probe and a SYBR Green I based real-time PCR assay were developed to detect and quantify Cladosporium in aerosols. The two real-time PCR systems proved to be highly specific and sensitive for Cladosporium. These methods were employed to quantify Cladosporium in aerosols of five different indoor environments. High spore concentration of Cladosporium (107/m3) was observed in a cow barn. Cladosporium spore concentration in paper and pulp factory and countryside house also exceeded threshold value for clinical significance. Prolonged exposure in these environments could impose certain health risk. Thus, monitoring Cladosporium spore concentration in indoor environments is important for indoor air quality control.

DNR žymenys yra polimorfinių DNR lokusų aleliai, lokalizuoti tam tikrose genomo vietose ir nustatomi naudojant įvairius molekulinės biologijos metodus. Genetinės įvairovės nustatymas ir tyrimas naudojant DNR žymenis gali padėti suprasti daugelio biologinių reiškinių, vykstančių augaluose (ląstelės, organizmo, rūšies lygmenyje), prigimtį. Vykdydami aptariamus tyrimus norime rasti atsakymą į pagrindinį klausimą: kokios naudotų DNR žymenų galimybės kai kurioms augalų biologijos problemoms (genetinių išteklių išsaugojimo, adaptyvumo, populiacijų diferenciacijos, biotinio ir abiotinio streso ir kt.) spręsti? Naudodami įvairius DNR žymenis kaip molekulinius instrumentus, įvertinome genetinės įvairovės lygį pagrindinėse miežių veislėse, sukurtose Baltijos šalyse ir Baltarusijoje, pritaikėme DNR žymenų nustatymo metodus Rubus idaeus, Lonicera ceruleae genetinėse kolekcijose esančių pavyzdžių genotipavimui bei miško medžių (Pinus sylvestris, Piceae abies) rinktinių medžių klonų tapatumo nustatymui; naudodami EST žymenis klonavome ir ištyrėme du genus, dalyvaujančius Solanum tuberosum atsake į biotinį (Erwinia carotovora infekcija) ir abiotinį stresą. Naudodami RAPD metodą ištyrėme Saxifraga hirculus, Piceae abies, Fraxinus excelsior, Taxus baccata, Rubus idaeus, Pinus sylvestris populiacijų genetinę struktūrą, diferenciacijos lygį, kai kurių ekologinių veiksnių įtaką genetinės įvairovės pasiskirstymui populiacijose. / DNA markers are alleles of polymorphic DNA loci that are established using methods of molecular biology and can be used for the identification of specific chromosome region. DNA markers are applied for the detection and analysis of genetic variation. These molecular instruments can help in the understanding of molecular basis of various biological phenomena in plants (loss of genetic diversity, population divergence, adaptivity, response to biotic and abiotic stress, genetic instability et cetera). The use of DNA markers in practical studies requires a careful consideration of the advantages and as well as limitations of various marker techniques. In this review various applications of DNA markers in plant genetic studies including genotyping and characterization of accessions of germplasm collections, assessment of genetic relationships between cultivars, understanding of the genetic variation within and between populations, plant genome analysis and gene cloning are discussed.

Magistro darbe lyginama vieno paprastosios pušies (Pinus sylvestris L.) klono skirtingų lajos dalių sėklinių palikuonių, augančių Višakio Rūdos bandomuosiuose želdiniuose, genetinė įvairovė remiantis DNR žymenimis ir kokybiniais bei kiekybiniais požymiais. Darbo objektas – paprastosios pušies (Pinus sylvestris L.) sėklinių palikuonių želdiniai iš skirtingų lajos dalies sėklų (viršutinės, vidurinės, apatinės). Darbo metodai – genetinė įvairovė tirta atliekant želdinių kiekybinių ir kokybinių požymių analizę bei jų DNR polimorfizmo tyrimą. Darbo rezultatai. Fenotipinių požymių tyrimai parodė, kad 24 m amžiuje, viršutinės lajos dalies sėkliniai palikuonys pasižymi ženkliai didesniu išlikimu, bet esminiai nesiskiria savo kiekybinių ir kokybinių požymių įvairove nuo apatinės lajos dalies palikuonių. Gali būti, kad fenotipinių požymių įvairovei atsiskleisti trukdo nevienodas medžių išlikimas, kur esant mažesniam išlikimui, susidaro nevienodi tarpai tarp medžių, galėję įtakoti didesnę erdvinę įvairovę radialiniam prieaugiui. 6 chloroplasto DNR(cpSSR) lokusų DNR polimorfizmo tyrimai parodė, kad viršutinės lajos dalies sėklinių palikuonių genetinė įvairovė yra ženkliai didesnė nei vidurines ir apatinės lajos dalių sėklinių palikuonių. Iš 30 skirtingų cpSSR tipų (tėvinių genotipų) 18 buvo aptikti viršutinės lajos dalies palikuonyse, tuo tarpu vidurinėje dalyje – 8, o apatinėje - 10 .Visi 5 proc. savidulkinių individų aptikti tarp apatinės ir centrinės lajos dalies palikuonių (18 vnt.)... [toliau žr. visą tekstą] / The genetic diversity of the progeny from different parts of Scots pine crown of a single clone by quantitative and qualitative traits and DNA polymorphism was investigated in the work of master science. Object of the work - 24 years old progeny from different parts of Scots pine crown of a single clone. The aim of the study - to compare the diversity of quantitative and qualitative traits and DNA polymorphism of24 years old progeny from different parts of Scots pine crown of a single clone by using cpSSR DNA markers. Methods of the work - Survival, stem diameter, stem straightness, flowering, cone yield, barktype,condition,the beginning of activegrowth and other parameters of the seedling progenies were evaluated. The genetic diversity was assessed at six cpDNA loci by the aid of cpSSRs. Study results. The results showed, that the survival of the progeny from the middle and the bottom of the crown was much lower than from the top. However, there were not any significant differences nor in the other traits neither between the variances of the progeny from the different parts of the crown. A reason could be that owing to low survival of the bottom and middle progeny, the remaining trees grew in a wider spacing and this uneven spacing between the treatments disturbed the comparison. cpSSR markers revealed much greater haplotype and allele diversity of the progeny from the top of the crown. Selfing rate was 5; background pollination 50; as much as 28 of the progeny from the... [to full text]

Made available in DSpace on 2016-01-26T18:56:21Z (GMT). No. of bitstreams: 1 Dissertacao.pdf: 378123 bytes, checksum: 7a0355c2b9d1cf6a5f844f5bb5622e1d (MD5) Previous issue date: 2008-12-01 / Cattleya walkeriana is widely grown by collectors and receive special attention by the breeders because it exhibit a rare colour variation and a great visual effect. RAPD markers (Random Amplified Polymorphic DNA) are one of the molecular markers most used in genetic studies. They have advantages, such as low amount of genomic DNA, hybridization absence, gene amplification at low costs and easy implementation. The DNA extracted from Cattleya walkeriana plants, wild and from cerulean offsprings, was amplified by elected RAPD primers. With the generated bands it was possible to estimate the fixation allelic index (FST) and to construct a dendogram. It could be inferred that Cattleya walkeriana is an allogamous species and that was a decrease in the genetic variability among the crosses as the recessive plants were used. Cerulean plants are difficult to obtain since they have all the pairs of genes in the recessive form. The inheritance for the trait in question is oligogenic and determined by genes that act in a complementary manner and are segregated independently. / Cattleya walkeriana é amplamente cultivada por colecionadores e tem recebido especial atenção dos melhoristas por apresentar variações na forma da coloração que são raras de serem obtidas, e possuem grande valor visual. Marcadores RAPD (polimorfismo de DNA amplificado ao acaso) são um dos tipos de marcadores moleculares mais utilizados em estudos genéticos por apresentarem vantagens como a utilização de baixa quantidade de DNA genômico, ausência de hibridação, emprego de amplificação gênica além de baixo custo e fácil execução. Para a realização deste trabalho, foi extraído DNA de diversas plantas de Cattleya walkeriana, plantas nativas e de progênies, obtidas de outras plantas de forma cerúlea, cujo DNA foi amplificado por marcadores RAPD selecionados. A partir das bandas geradas foi possível estimar o índice de fixação alélica (FST) e construir um dendrograma das amostras. Determinou-se que Cattleya walkeriana é uma espécie alógama e que houve diminuição da variabilidade genética dentro dos cruzamentos á medida que plantas recessivas de mesmo fenótipo foram utilizadas. Plantas cerúleas são de difícil obtenção, pois apresentam todos os pares de genes na forma recessiva. A herança para a característica em questão é oligogênica e determinada por genes que atuam de maneira complementar e são de segregação independente.

Made available in DSpace on 2016-07-18T17:51:05Z (GMT). No. of bitstreams: 1 Dissertacao.pdf: 378123 bytes, checksum: 7a0355c2b9d1cf6a5f844f5bb5622e1d (MD5) Previous issue date: 2008-12-01 / Cattleya walkeriana is widely grown by collectors and receive special attention by the breeders because it exhibit a rare colour variation and a great visual effect. RAPD markers (Random Amplified Polymorphic DNA) are one of the molecular markers most used in genetic studies. They have advantages, such as low amount of genomic DNA, hybridization absence, gene amplification at low costs and easy implementation. The DNA extracted from Cattleya walkeriana plants, wild and from cerulean offsprings, was amplified by elected RAPD primers. With the generated bands it was possible to estimate the fixation allelic index (FST) and to construct a dendogram. It could be inferred that Cattleya walkeriana is an allogamous species and that was a decrease in the genetic variability among the crosses as the recessive plants were used. Cerulean plants are difficult to obtain since they have all the pairs of genes in the recessive form. The inheritance for the trait in question is oligogenic and determined by genes that act in a complementary manner and are segregated independently. / Cattleya walkeriana é amplamente cultivada por colecionadores e tem recebido especial atenção dos melhoristas por apresentar variações na forma da coloração que são raras de serem obtidas, e possuem grande valor visual. Marcadores RAPD (polimorfismo de DNA amplificado ao acaso) são um dos tipos de marcadores moleculares mais utilizados em estudos genéticos por apresentarem vantagens como a utilização de baixa quantidade de DNA genômico, ausência de hibridação, emprego de amplificação gênica além de baixo custo e fácil execução. Para a realização deste trabalho, foi extraído DNA de diversas plantas de Cattleya walkeriana, plantas nativas e de progênies, obtidas de outras plantas de forma cerúlea, cujo DNA foi amplificado por marcadores RAPD selecionados. A partir das bandas geradas foi possível estimar o índice de fixação alélica (FST) e construir um dendrograma das amostras. Determinou-se que Cattleya walkeriana é uma espécie alógama e que houve diminuição da variabilidade genética dentro dos cruzamentos á medida que plantas recessivas de mesmo fenótipo foram utilizadas. Plantas cerúleas são de difícil obtenção, pois apresentam todos os pares de genes na forma recessiva. A herança para a característica em questão é oligogênica e determinada por genes que atuam de maneira complementar e são de segregação independente.

Central to the application of genomics to animal agriculture are DNA markers, especially microsatellites and single nucleotide polymorphisms. These markers are the resources necessary for constructing genetic maps and for determining how improved and unimproved animal breeds are related. Here, DNA markers were developed for two avian species, the turkey, Meleagris gallopavo, and the budgerigar (budgie), Melopsittacus undulatus. Genomic libraries enriched for simple sequence repeats were used to generate about 70 budgie sequences of a total length of 38 kb. From these sequences, 9 primer pairs were designed and used to screen for informativeness in a panel of DNA samples from unrelated budgie samples. All but one of the nine primers evaluated were polymorphic with the number of alleles ranging from two to four. Comparative analysis involving the use of these budgie primers showed moderate sequence similarity to turkey and chicken. The genomic libraries and the comparative sequences provide useful genomic reagents that could be used to construct a budgie genome map. In the turkey, ten previously described microsatellites and a gene-based single nucleotide polymorphism (SNP) were used to evaluate the relatedness of heritage varieties to a commercial strain. Estimates of Nei's genetic distance (D) and genetic differentiation (Rst) between populations using microsatellite markers showed that the commercial strain is genetically more closely related to the Bourbon Red and Narragansett and least related to the Royal palm and Spanish Black. Gene flow (Nm) level was highest between the commercial and Bourbon Red populations. The SNP analysis by PCR-RFLP revealed that the commercial strain was more closely related to the Spanish black and Narragansett and least related to the Bourbon red and Blue slate. Though results of the two marker systems, microsatellite and SNP, were inconsistent, they provide insights into using heritage turkeys to genetically improve commercial populations by introgression. The present thesis investigation showed that DNA markers provide a strong opportunity to develop genomic reagents needed to test hypotheses in little-studied agriculturally important and model avian species. / Master of Science

The objective of this study was to identify DNA markers for each of six added C-genome chromosomes in Triticum aestivum L. cv. 'Alceso'-Aegilops caudata L. addition lines using the randomly amplified polymorphic DNA (RAPD) technique. DNA from Ae. caudata, T. aestivum, amphiploid of T. aestivum X Ae. caudata, and six disomic addition lines of wheat having a pair of Ae. caudata chromosomes was used as the template for the amplification of RAPD markers with a total of 58 random 10-mer oligonucleotide primers. Two primers, OPC-08 and OPJ-16, produced one intense band each from the amphiploid of T. aestivum X Ae. caudata and Ae. caudata, which was absent in all six addition lines. Each of these two primers produced a chromosome marker that could be tentatively located to the chromosome CA of Ae. caudata. OPJ-02, OPD-12, OPD-02, OPJ-12, OPD-20, and OPJ-14 produced a marker each for CB, CC, CD, CE, CF, and CG, respectively. OPJ-09 produced C-genome chromosome-specific RAPD markers. Also, OPC-05 and OPJ-19 produced RAPDs from both wheat and Ae. caudata genomes.

Made available in DSpace on 2016-08-12T19:15:25Z (GMT). No. of bitstreams: 1 leidianeBA_DISSERT.pdf: 951596 bytes, checksum: 769c70c85ea3d6f0353cabbe1093e973 (MD5) Previous issue date: 2014-02-28 / Coordenação de Aperfeiçoamento de Pessoal de Nível Superior / Downy mildew, caused by the fungus Pseudoperonospora cubensis, is a major threat to the production of melons in wetlands worldwide. Despite its importance, there are few studies involving the assessment of sources of resistance, as well as the identification of molecular markers that are connected to this feature and that may help breeders in marker-assisted selection (MAS). Given this, the objectives of this work were promote the selection of sources of resistance to downy mildew, from melon accessions, collected from different states in the Northeast region of Brazil and to identify QTLs that co-segregate with the resistance gene of this pathogen, using the F2:3 obtained from parents and Védrantais MR-1, contrasting to the register character. To identify sources of resistance, thirty-six accesses and four commercial cultivars in a randomized block design (RBD) were assessed with three replications and seven plants per plot, which evaluation was made on the field. The evaluation was carried out on 24 days after transplanting the seedlings to the field, when 50% of plants reached the stage of flowering, with the aid of a diagrammatic scale. It was observed that there is variability in melon germplasm for reaction to downy mildew. The accessions A1, A4, A6, A9, A12, A17, A18, A23, A27, A35 and A36 were considered promising for use in breeding programs aiming resistance to P cubensis. All commercial cultivars analyzed were highly susceptible to the pathogen. To identify markers connected the resistance QTLs, MR-1 and Védrantais parents were crossed and subsequently the F2 generation was obtained. It was extracted the DNA for evaluation molecular of the 98 plants obtained with microsatellite primers polymorphic selected from contrasting parents. Of F2 plants were obtained 98 progenies F2:3 that were used for the assessments phenotypic of the severity caused by P. cubensis, it was used the same time assessment described in the previous experiment, the period April-June 2013. The experimental design utilized was lattice simple 10 x 10. The efficiency of lattice was low (100.28%) thus data were analyzed in DBC. Significant genetic differences (P<0.01) between the progenies were identified. The estimated of heritability was considered high (86.75%), indicating successful selection. Were found 23 primers polymorphic, of this sum, thirteen amplified in all the population and were used for analyzes genotypic. Of these, ten markers presented values below level critical specified by FDR test. There was more proportion allele of parent Védrantais for eight loci, while there was more proportion allele the parent MR-1 for two loci. For other loci (three), frequencies allelic did not change. The values coefficients of determination (R2) obtained for the markers were relatively low in population. By regression simple linear, the markers explained the feature in population F2:3 the most were CMBR 139 and CMMS 22-2, being the same identified by regression multiple linear, thus they can also be used in SAM / O míldio, causado pelo fungo Pseudoperonospora cubensis, é uma das principais ameaças à produção do meloeiro em áreas úmidas em todo o mundo. Apesar de sua importância, são poucos os trabalhos envolvendo a avaliação de fontes de resistência, bem como a identificação de marcadores moleculares que estejam ligados a esta característica e que possam auxiliar os melhoristas na seleção assistida por marcadores (SAM). Diante disso, os objetivos deste trabalho foram promover a seleção de fontes de resistência ao míldio, a partir de acessos de meloeiro, coletados em diferentes estados da região Nordeste do Brasil e também identificar QTLs que co-segreguem com genes de resistência a este patógeno, utilizando progênies F2:3 obtidas dos genitores MR-1 e Védrantais, contrastantes para o caráter. Para a identificação de fontes de resistência, foram avaliados trinta e seis acessos e quatro cultivares comerciais em delineamento em blocos casualizados (DBC) com três repetições e sete plantas por parcela, cuja avaliação foi feita em campo. A avaliação se deu aos 24 dias após o transplantio das mudas para o campo, depois que 50% das plantas atingiram o estágio de floração, com o auxílio de uma escala diagramática. Observou-se que existe variabilidade no germoplasma de meloeiro para reação ao míldio. Os acessos A1, A4, A6, A9, A12, A17, A18, A23, A27, A35 e A36 foram considerados promissores para o uso em programas de melhoramento visando resistência a P cubensis. Todas as cultivares comerciais analisadas foram altamente suscetíveis ao patógeno. Para identificação de marcadores ligados a QTLs de resistência, os genitores MR-1 e Védrantais foram cruzados e posteriormente foi obtida a geração F2. Das 98 plantas obtidas foi extraído o DNA para avaliação molecular com primers microssatélites polimórficos selecionados a partir dos genitores contrastantes. Das plantas F2 foram obtidas 98 progênies F2:3 que foram utilizadas para as avaliações fenotípicas quanto a severidade provocada por P. cubensis, seguindo a mesma época de avaliação descrita no experimento anterior no período de abril a junho de 2013. O delineamento utilizado foi o látice simples 10 x 10. A eficiência do látice foi baixa (100,28%), por isso, os dados foram analisados em DBC. Foram identificadas diferenças genéticas significativas (P<0,01) entre as progênies. A estimativa de herdabilidade foi considerada alta (86,75%), indicando sucesso com a seleção. Foram encontrados 23 primers polimórficos. Treze amplificaram em toda a população e foram utilizados para as análises genotípicas. Destes, dez marcadores apresentaram valores abaixo do nível crítico especificado pelo teste FDR. Para oito locos houve maior proporção de alelos do genitor Védrantais, enquanto que para dois locos houve maior proporção de alelos do genitor MR-1. Para os demais locos (três), as frequências alélicas não se alteraram. Os valores dos coeficientes de determinação (R2) obtidos para os marcadores foram relativamente baixos na população. Pela regressão linear simples, os marcadores que mais explicaram a característica na população F2:3 foram CMBR 139 e CMMS 22-2, sendo os mesmos identificados pela regressão linear múltipla, podendo, portanto, serem utilizados na SAM