It is routine for the forensic scientist to obtain a genetic profile of an individual from DNA recovered from a biological stain deposited at a crime scene. In contrast, only a limited number of laboratories in the United States have the capability of performing Y-STR analysis in casework. In order to aid in facilitating the transfer of Y-STR technology to the crime laboratory community for operational use, a comparison between commercial products from three main vendors (Applied Biosystems AmpFLSTR Yfiler PCR Amplification Kit, Promega PowerPlex - Y System, Reliagene Y-PLEX 12) and two in-house Y-STR multiplexes (MPI and MPB) commenced. The main intention for this comparison was to ascertain whether commercial Y-STR kits are able to obtain a male profile from difficult samples which have been accomplished with our in-house Y-STR multiplexes; such as mixtures, post coital specimens, and environmental insults. To aid the crime laboratory community an in depth comparison of the three main commercial Y-STR kits began in hopes to glean information in circumstances where Y chromosome polymorphisms may need to be employed. For example, the ability to provide investigators with the numbers of semen donors in multiple rape cases, identification of the genetic profile of the male component in a male/female mixture, and identification of the genetic profile of the male component in an extended interval post-coital sample. The capability of typing Y-STR loci by the crime laboratory community could dramatically affect the admissibility of Y-STR evidence. Therefore, the comparison of commercially available kits is an imperative process by which the scientific community acquires the necessary information to assess the ability of a procedure to obtain reliable results, determine the conditions under which such results can be obtained and define the limitations of the procedure. Thus the information for the study could lend itself to a standard being established amongst Y-STR kits for operational use and/or the production of a new Y-STR kit. One example of how the comparison of the three main commercial Y-STR kits could directly impact a new standard being established is by examining post-coital samples and their extreme limits (>48 hrs) for each kit in which a full male genetic profile was observed and comparing it to other commercial Y-STR kit and in-house Y-STR multiplexes. This would help establish the types of cases where specific Y-STR kits would be most useful, and the parameters in which each kit is able to perform. Thus leading to the development of a highly sensitive Y-STR kit that would be more sufficient to perform with the variety of samples an operational crime laboratory would routinely analyze. The capability of typing Y-STR loci by the crime laboratory community could dramatically affect the admissibility of Y-STR evidence. Therefore, the comparison of commercially available kits is an imperative process in order to inform the forensic community of different Y-STR kits available and their performance through direct comparison using modified SWGDAM validation guidelines.

Discovering, screening, and associating changes in DNA sequence are important to a broad range of disciplines and play a central role in Forensic Science. The typical types of changes include sequence variations [single nucleotide polymorphisms (SNP)] and length variations [short tandem repeats (STR)]. The steps for forensic DNA sample processing are similar for both types of changes but diverge at the point of detection. A number of approaches are being explored for SNP genotyping while STR analysis primarily consists of size-based analysis by capillary electrophoresis. Limitations exist for all current detection methods that pose significant impacts to forensic analysis. Bi-allelic SNPs result in three possible genotypes with a minimal amount of information generated per marker. Limitations for SNP analysis are due to the inability to amplify a suitable number of SNP markers from low DNA content samples to provide an appropriate level of discrimination. Multi-allelic STR markers are currently the marker of choice for forensic typing but a variety of experimental artifacts are possible that consist of either biology or technology related causes. Molecular genotyping methods developed across other disciplines have potential to alleviate some of these shortcomings but no current approach is capable of genotyping both SNP and STR loci with a single chemistry. The need for a more effective, efficient, and generalized approach led to development of a unique method called Dye Probe Fluorescence Resonance Energy Transfer (dpFRET) and determination of its suitability for forensic analysis. The development phase of the research consisted of synthetic testing to establish proof of concept for the chemistry followed by polymerase chain reaction (PCR) based assays to demonstrate real world applications. Following successful development, the boundaries and limitations for the technology were established (sensitivity, allelic dropout, mixed samples) and efforts were made to improve the approach. In the process, parallel testing for other fields including molecular pathology and conservation biology were incorporated to explore potential widespread application of this new approach. The overall goal of this project was to develop and explore the limitations for a unique approach to genotyping both SNPs and STRs. A majority of the work involved development of the method itself with the ultimate objective of application for forensic science. The focus of this project was to address and alleviate some of the shortcomings of current approaches that result in potential limitations for forensic analysis. It is expected that future applications of this technology might impact a wide range of disciplines to aid in discovery, screening and association of changes in DNA sequence.

No description available.

Made available in DSpace on 2016-08-10T10:38:27Z (GMT). No. of bitstreams: 1 Lindomar Valentin Ferreira.pdf: 1215853 bytes, checksum: 2cb0836a827c452d5ca06c97f0d59f35 (MD5) Previous issue date: 2011-05-13 / The population genetics aims to study the allelic and genotypic frequencies in populations and the mechanisms that these frequencies change over generations. This study aims to estimate and compare the allele frequencies of 15 autosomal STR markers, sheets of 501 tests the genetic link in 986 individuals in the population of Goiânia-Goiás. The markers that showed higher numbers of alleles were D18S51 and Penta E with 20 to 19. The marker that had allele frequency was higher for the TPOX 8 allele. Other statistical parameters analyzed demonstrated high PIC, PD and PE for the marker Penta E. Analysis of allele frequencies obtained in this study were statistically compared with populations of five regions of Brazil, by Student's t test, not intra-population differences were observed (p> 0.05). In this context, the genetic database based on the values of the allele frequencies of the population of Goiânia, can be used in testing the genetic link, as well as in studies of human identification, due to the similarities observed in other Brazilian populations. / A genética de populações visa o estudo das frequências alélicas e genotípicas em populações e os mecanismos capazes de mudar estas frequências ao longo das gerações. Essa pesquisa tem como objetivo estimar e comparar as frequências alélicas de 15 marcadores autossômicos STR, de 501 planilhas de exames de vínculo genético em 986 indivíduos da população de Goiânia-Goiás. Os marcadores que apresentaram maiores números de alelos foram o D18S51 com 20 e o Penta E com 19. O marcador que teve maior frequência alélica foi o TPOX para o alelo 8. Outros parâmetros estatísticos analisados caracterizaram elevados PIC, PD e PE para o marcador Penta E. As análises das frequências alélicas obtidas neste estudo foram comparadas estatisticamente com populações de cinco regiões brasileiras, pelo teste t Student, e não foram verificadas diferenças intra populacionais significativas (p>0,05). Neste contexto, o banco de dados genéticos baseado nos valores das frequências alélicas da população de Goiânia, pode ser utilizado em testes de vínculo genético, assim como em estudos de identificação humana, devido às semelhanças verificadas com outras populações brasileiras.

Made available in DSpace on 2016-08-10T10:38:39Z (GMT). No. of bitstreams: 1 Ian Marques Candido.pdf: 1079357 bytes, checksum: dac0a34c413d6875a597e0f2ba2a41f6 (MD5) Previous issue date: 2013-02-08 / The identification of human remains in decomposition, charred, skeletal remains and mass disasters can be performed by Forensic Genetics and, in most cases, bones and teeth are the only viable source for DNA typing .Thus, considering the large number of bones used in human identification and the need for standardization of DNA extraction in this kind of sample, the aim of this study is to compare two techniques of DNA extraction, with the possibility of automation. The analysis was performed on twenty five human bones evaluating the quantity of the extracted genetic material, genetic profiles obtained for each sample and the time analysis by method used. With magnetic bead in platform automated, analysis time was 3 hours to process 12 samples, whereas by silica column four samples in 27 hours. Magnetic bead recovered a larger amount of DNA in 88% of samples. 68% of the samples magnetic particle had a high amplification partial (9/16 loci) and silica column only 36%. Therefore, the method used magnetic bead is suitable for automating the extraction processes. / A identificação humana de restos mortais em avançado estado de decomposição, carbonizados, desastres em massa e esqueletizados pode ser realizada pela Genética Forense e, na maioria das vezes, ossos e dentes são as únicas fontes de DNA viáveis para análise. Dessa maneira, considerando o grande número de ossos utilizados na identificação humana e a necessidade de padronização da técnica de extração de DNA nesse tipo de amostra, o objetivo do presente trabalho é comparar duas técnicas de extração de DNA, com possibilidade de automação. A análise foi realizada em vinte e cinco ossos humanos avaliando a quantidade do material genético extraído, os perfis genéticos obtidos em cada amostra e o tempo de análise gasto pela metodologia de extração. Com a metodologia de extração por partículas magnéticas utilizando plataforma automatizada, o tempo de análise foi de 3 horas para processar 12 amostras, enquanto que por coluna de sílica 4 amostras em 27 horas. Partícula magnética recuperou uma maior quantidade de DNA em 88% das amostras. 68% das amostras extraídas por partículas magnéticas tiveram uma amplificação parcial alta (9/16 loci) e por coluna de sílica apenas 36%. Por conseguinte, a metodologia de extração por partículas magnéticas é apropriada para a automatização dos processos de extração.

Made available in DSpace on 2016-08-10T10:39:19Z (GMT). No. of bitstreams: 1 BRAULIO CANCADO FLORES.pdf: 638020 bytes, checksum: 0d45d0c26c601b5579ff1ec8f22426fc (MD5) Previous issue date: 2008-06-27 / The most serious radiological accident occurred in the western hemisphere and it happened in September of 1987, when a radiotherapy unit containing 50.9 TBq of Cs137Cl was removed from an abandoned radiotherapy clinic in the State of Goias, Brazil and it was subsequently disassembled. This event provides an opportunity to assess the genetic effects of ionizing radiation. We investigated the genetic variation of 12 microsatellite loci in 11 families of exposed liquidators, who worked at the civil defense procedures. As a control group, non-exposed families, comprised of 900 individuals, from the same area of Goias state were analyzed. When we compared the exposed and control groups, we found an increase in the number of new alleles in the offspring of the exposed liquidators. The mutation rate was found to be higher in the exposed families compared to the control group. These results, associated with previous studies, indicated that exposure to ionizing radiation can be detected in offspring of exposed liquidators and also suggest that the elevated microsatellite mutation rate can be attributed to radioactive exposure. / O acidente radiológico mais grave do ocidente aconteceu em setembro de 1987, quando um aparelho de radioterapia contendo 50.9 TBq de Cs137Cl foi removida das ruínas de uma clínica de radioterapia e em seguida, violada. Este acontecimento criou a oportunidade de que se investigasse os efeitos genéticos da radiação ionizante. Foi investigada a variabilidade genética em 12 loci microssatélites em 11 famílias de liquidators, que trabalharam nas ações de defesa civil. Como grupo controle, utilizamos famílias sem potencial de exposição à radiação ionizante, perfazendo 900 indivíduos analisados, da mesma área do Estado de Goiás. Quando comparamos os grupos exposto e controle, encontramos um incremento no número de novos alelos nos perfis dos liquidators expostos. A taxa de mutação encontrada foi maior nas famílias dos expostos do que no grupo controle. Estes resultados, associados a anteriores, indicaram que a análise dos perfis alélicos dos liquidators pode indicar a exposição à radiação ionizante, além de uma elevada taxa de mutações de microssatélites poder ser atribuída a esta exposição

L'étude des parentés génétiques permet à l'anthropologie d'identifier la place du sujet au sein des différentes structures dans lesquelles il évolue : l'individu est membre d'une famille biologique, d'un groupe social et d'une population. L'application des méthodes probabilistes classiques (établies pour répondre à des problématiques de médecine légale, comme la méthode des Likelihood Ratios (LR) ou " Rapports de vraisemblance ") aux données STR issues du matériel archéologique a permis la découverte de nombreux liens de parenté, qui ensemble constituent des généalogies parfois complexes. Notre pratique prolongée de ces méthodes nous a cependant amenés à identifier certaines limites de l'interprétation des données STR, en particulier dans les cas de parentés complexes, distantes ou consanguines, ou dans des populations isolées, méconnues ou disparues. Ce travail de thèse s'attache en premier lieu à quantifier la fiabilité et l'efficacité de la méthode des LR dans quatre situations : une population moderne avec une grande diversité allélique, une population moderne avec une faible diversité allélique, une population ancienne de grande taille et une population ancienne de petite taille. Les publications récentes font usage des marqueurs plus nombreux issus des nouvelles technologies de séquençage (NGS) pour mettre en place de nouvelles stratégies de détection des parentés, basées en particulier sur l'analyse des segments chromosomiques partagés par ascendance entre les individus (segments IBD). Ces méthodes ont rendu possible l'estimation plus fiable de probabilités de parenté dans le matériel ancien. Elles sont néanmoins inadaptées à certaines situations caractéristiques de la génétique des parentés archéologiques : elles ne sont pas conçues pour fonctionner avec une seule paire isolée d'individus et reposent, comme les méthodes classiques, sur l'estimation de la diversité allélique dans la population. Nous proposons donc une quantification de la fiabilité et de l'efficacité de la méthode des segments partagés à partir de données NGS, en s'attachant à déterminer la qualité des résultats dans les différentes situations qui correspondent à des tailles de population plus ou moins importantes et à une hétérogénéité plus ou moins grande de l'échantillonnage.[...] / The study of genetic kinship allows anthropology to identify the place of an individual within which they evolve: a biological family, a social group, a population. The application of classical probabilistic methods (that were established to solve cases in legal medicine, such as Likelihood Ratios, or LR) to STR data from archaeological material has permitted the discovery of numerous parental links which together constitute genealogies both simple and complex. Our continued practice of these methods has however led us to identify limits to the interpretation of STR data, especially in cases of complex, distant or inbred kinship. The first part of the present work is constituted by the estimation of the reliability and the efficacy of the LR method in four situations: a large modern population with significant allelic diversity, a large modern population with poor allelic diversity, a large ancient population and a small ancient population. Recent publications use the more numerous markers analysed using Next generation Sequencing (NGS) to implement new strategies in the detection of kinship, especially based on the analysis of chromosome segments shared due to common ancestry (IBD "Identity-by-Descent" segments). These methods have permitted the more reliable estimation of kinship probabilities in ancient material. They are nevertheless ill-suited to certain typical situations that are characteristic of ancient DNA studies: they were not conceived to function using single pairs of isolated individuals and they depend, like classical methods, on the estimation of allelic diversity in the population. We therefore propose the quantification of the reliability and efficiency of the IBD segment method using NGS data, focusing on the estimation of the quality of results in different situations with populations of different sizes and different sets of more or less heterogeneous samples.[...]

Au sein de la Sibérie Orientale, l'origine des Yakoutes reste une énigme qui demeure non résolue par les études classiques. Les Yakoutes représentent en effet le seul peuple d'éleveurs de bétail et de chevaux au sein d'un ensemble de populations composé de chasseurs et d'éleveurs de rennes. Leur langue, mélange de mots d'origine turque et mongole, ainsi que leurs pratiques culturelles accentuent encore ce contraste avec les populations alentour. <br />L'analyse moléculaire de spécimens yakoutes anciens apparaissait comme bien adaptée à l'étude de la formation de ce peuple. En effet, l'évolution récente des techniques de biologie moléculaire rend aujourd'hui possible l'analyse génétique des populations du passé et les conditions environnementales rencontrées en Sibérie Orientale sont particulièrement propices à la conservation des acides nucléiques. <br />L'étude de plus de 60 sujets anciens provenant de Yakoutie Centrale, a permis d'obtenir des résultats originaux concernant, d'une part, l'aspect moléculaire de ce travail et d'autre part, l'ethnogenèse yakoute. Ces résultats ont notamment permis de souligner la très grande qualité des échantillons provenant de Sibérie Orientale. Il a ainsi été possible d'étudier des marqueurs génétiques rarement analysables dans les études d'anthropologie moléculaire, comme les STR autosomaux et du chromosome Y, et d'obtenir des résultats uniques sur des substrats difficiles. Cet ensemble de facteurs a conduit à l'obtention de données dont l'authenticité est manifeste. En outre, le nombre important de sujets étudiés ainsi que la mise en parallèle de nouveaux protocoles sur plusieurs types de prélèvements ont permis d'apporter des informations sur les propriétés de ces différents substrats. <br />La comparaison des résultats collectés pour le chromosome Y et l'ADN mitochondrial des sujets anciens avec les populations voisines mais également du sud de la Sibérie, ont permis de dégager de nouvelles hypothèses concernant les origines des lignées paternelles et maternelles des Yakoutes. L'influence méridionale semble confirmée par notre étude, mais nos résultats mettent également en avant que des contacts anciens, précédant les migrations déjà proposées, ont du se produire entre les peuples nomades des steppes et les populations de Sibérie Orientale. De plus, la population yakoute apparaît extrêmement stable au cours des siècles malgré des changements notables dans les influences culturelles et l'arrivée des colons russes au XVIIIième siècle. Ainsi, certaines lignées masculines, spécifiques à la population yakoute, montrent une pérennité exceptionnelle puisqu'elles se sont maintenues depuis le XVième siècle jusqu'à nos jours, avec des fréquences très importantes.<br />La qualité des données obtenues ainsi que les conclusions qui ont pu être proposées suite à ce travail de recherche confirment à nouveau la pertinence de la mise en œuvre d'un approche moléculaire dans la compréhension de la formation et de l'organisation des populations du passé.

O uso de marcadores do tipo STR e SNP tem se revelado de grande importância na discriminação entre indivíduos de uma mesma população, assim como para estudos evolutivos. A utilização de um conjunto de 17 STRs e 46 SNPs específicos de cromossomo Y permitiu a caracterização de um conjunto de amostras representativas das populações do Rio de Janeiro e do oeste africano, com uma avaliação mais ampla sobre a ancestralidade de origem paterna. Na primeira parte deste estudo foram analisados 605 indivíduos do sexo masculino do estado do Rio de Janeiro. Como resultado, não foram observadas diferenças significativas entre as populações do sudeste e do Rio de Janeiro, que apresentou uma alta diversidade de haplótipos (0,9999 0,0001) e de haplogrupos (0,7589 0,0171). A comparação da população miscigenada do Rio de Janeiro com diferentes grupos étnicos ou populacionais mostrou que a frequência de indivíduos com marcadores tipicamente Europeus é de 77%, africanos é de 14,87% e em ameríndios é de 2,31%. A segunda parte do estudo revelou uma grande diversidade haplotípica (1,0000 0,0018) numa amostra do Oeste africano. Quanto ao valor da diversidade de haplogrupos (0,6895 0,0200), este foi similar aos observados em populações de origem Bantu do oeste e centro africanos, principalmente de Benin, Nigéria e Costa do Marfim. A terceira parte deste estudo mostrou que não existem diferenças significativas entre o componente africano da amostra do Rio de Janeiro e as populações africanas do sudeste, oeste e centro oeste. Por outro lado, observamos diferenças significativas quando comparamos o componente africano do Rio de Janeiro e o oeste africano com populações de Uganda, Quênia e África do Sul. A ampliação de estudos genéticos nas populações da África se fazem necessários para o entendimento da diversidade genética no mundo. Este trabalho contribuiu para fornecer mais alguns dados genéticos, que podem ser somados aos estudos mundiais que estão sendo realizados, ampliando os nossos conhecimentos sobre a formação das populações que também foram influenciadas pelo fenômeno da Diáspora Africana. / The use of STR and SNP markers has proved to be of great importance to discriminate individuals of the same population as well as for evolutionary studies. The use of a set of 17 STRs and 46 SNPs specific from the Y chromosome allowed the characterization of a group of samples representative of the Rio de Janeiro and the West African populations, with a deep assessment of the paternal ancestry. The first part of this study focused in the analysis of 605 males of the state of Rio de Janeiro. The results showed no significant differences between the Brazilian southeastern populations and Rio de Janeiro, which showed high values of haplotype (0.9999 0.0001) and haplogroup (0.7589 0.0171) diversities. The second part of the study revealed a high haplotype diversity (1.0000 0.0018) in a sample from West Africa. The value of haplogroup diversity (0.6895 0.0200) was similar to those previously seen in the West and Center African Bantu populations, mainly from Benin, Nigeria and Ivory Coast. The third part of this study showed no significant differences between the African component of our sample from Rio de Janeiro and the Southeastern, Western and Midwestern African populations. On the other hand, significant differences were observed when comparing the samples of the African component in Rio de Janeiro and of West Africa with population samples from Uganda, Kenya and South Africa. The increase of genetic studies in African populations is important for a better understanding of world genetic diversity. This work provided new genetic data, which can be added to those already available, expanding our knowledge about the formation of other populations also influenced by the African Diaspora.

Made available in DSpace on 2014-12-17T14:16:30Z (GMT). No. of bitstreams: 1 TaissaMMO_DISSERT.pdf: 1807409 bytes, checksum: 3f71ff29b01e86200d456b984c7e8b08 (MD5) Previous issue date: 2012-02-24 / Coordena??o de Aperfei?oamento de Pessoal de N?vel Superior / Human population have a significant number of polymorphic loci, whose use and applications range from construction of linkage maps, to study the evolution of populations, through the determination of paternity, forensic medicine and migration. Currently, STRs (Short Tanden Repeats) markers are considered the major markers for human identification, mainly due to its abundance and high variability because of the fact that they are easily amplifiable by PCR (Polymerase Chain Reaction), work with low amounts of DNA and be capable of automation processes involving fluorescence detection. The creation of regional databases containing allele frequencies of population provide subsidies to increase the reliability of the results of determining the genetic link. This paper aims to obtain a database of allele frequencies of 15 polymorphic molecular loci (D8S1179, D21S11, D7S820, CSF1PO, D19S433, vWA, TPOX, D18S51, D3S1358, TH01, D13S317, D16S539, D2S1338, D5S818 e FGA) in a population classifies as born in the State of Rio Grande do Norte, Brazil, totaling 1100 unrelated individuals. To evaluate the frequency, DNA samples were submitted to PCR amplification, followed by capilarry electrophoresis genetic sequencer. The frequencies identified in this study were compared with brazilian population in general and other states in Brazil. Except for the loci D21S11, D19S433 and D2D1338, the genotypes found were in Hardy-Weinberg equilibrium and no significant differences among the frequencies were found in the populations studied. The most informative loci was D2S1338 and D18S51, and the less informative is the locus TPOX / As popula??es humanas apresentam um consider?vel n?mero de loci polim?rficos, cujo uso e aplica??es v?o desde a constru??o de mapas de liga??o at? estudos de evolu??o das popula??es, passando pela determina??o de paternidade, medicina forense e migra??o. Atualmente, os STRs (Short Tandem Repeats) s?o considerados os marcadores de identifica??o humana por excel?ncia, t?tulo em grande parte devido a sua abund?ncia e elevada variabilidade, devido ao fato de serem facilmente amplific?veis pela Rea??o em Cadeia da Polimerase, PCR (Polymerase Chain Reaction), funcionarem com baixas quantidades de DNA e serem pass?veis de automa??o com processos envolvendo detec??o por fluoresc?ncia. A forma??o de bancos de dados regionais, contendo as frequ?ncias al?licas da popula??o de uma microrregi?o, fornece subs?dios para aumentar a confiabilidade dos resultados de determina??o de v?nculo gen?tico. Neste trabalho, visa-se a obten??o de um banco de dados das frequ?ncias al?licas de 15 loci polim?rficos moleculares (D8S1179, D21S11, D7S820, CSF1PO, D19S433, vWA, TPOX, D18S51, D3S1358, TH01, D13S317, D16S539, D2S1338, D5S818 e FGA), em uma popula??o classificada como nascida no Estado do Rio Grande do Norte, Brasil, totalizando 1100 indiv?duos n?o aparentados. Para avalia??o das frequ?ncias, amostras de DNA foram submetidas ? amplifica??o por PCR, e os gen?tipos foram obtidos atrav?s de eletroforese capilar em sequenciador gen?tico. As frequ?ncias apuradas no presente estudo foram comparadas com as da popula??o brasileira em geral e com as de outros estados do Brasil. Com exce??o dos loci D21S11, D19S433 e D2S1338, os gen?tipos encontrados da popula??o do RN mostram-se em Equil?brio de Hardy-Weinberg, e sem grandes diferen?as significativas entre as frequ?ncias encontradas nas popula??es estudadas. Os loci mais informativos foram D2S1338 e D18S51, enquanto que o locus menos informativo foi o TPOX