A região Neotropical é uma das regiões com maior biodiversidade no planeta. Essa região apresenta uma complexa história geológica que iniciou-se com a quebra da Gondwana, separando os continentes sul-americano e africano, durante o Mesozóico, há ca. de 150 milhões. Todas as mudanças geológicas que seguiram influenciaram a diversificação da fauna e flora dessa região. No entanto, ainda não sabemos quais processos levaram à alta diversidade encontrada nesta região. Esse estudo, foca em Stizophyllum, (Bignonieae, Bignoniaceae), um pequeno gênero de lianas Netropicais, que ocorre desde o México até o sul do Brasil. Apesar da cirscunscrição de Stizophyllum ser clara, limites específicos permanecem complicados neste grupo e pouco se sabe sobre sua história evolutiva. Este estudo visa: (i) reconstruir a filogenia do gênero e utilizá-la como base para inferir a história biogeográfica do grupo, e (ii) desenvolver marcadores de microssatélites (SSRs) nucleares e plastidiais, para futuros estudos filogeográficos e de genética de populações. Para tal, reconstruímos a filogenia do gênero com base em três marcadores moleculares (ndhF, rpl32-trn,L e pepC) e uma ampla amostragem de indivíduos, utilizando inferências bayesiana e de máxima verossimilhança. Em seguida estimamos as idades de divergência das diversas linhagens e reconstruímos a história biogeográfica do grupo. Por fim, desenvolvemos marcadores de microssatélite nucleares (nSSRs) e de cloroplasto (cpSSRs) para o grupo. Ao todo, desenvolvemos trinta e sete SSRs, nove nucleares e vinte oito de cloroplasto. Todos marcadores foram polimórficos em S. riparium e apresentaram sucesso de transferabilidade para S. inaequilaterum e S. perforatum. Cinco clados principais foram reconstruídos na filogenia molecular do gênero, os quais são caracterizados por bons caracteres morfológicos e aqui reconhecidos como espécies em uma nova sinopse apresentada para o grupo: (i) S. inaequilaterum Bureau & K. Schum., distribuído pela Amazônia e América Central; (ii) S. perforatum (Cham.) Miers, distribuído pela Mata Atlântica e Áreas Secas do Brasil Central, (iii) S. riparium (Kunth) Sandwith, distribuído por toda Bacia Amazônia; (iv) S. flos-ardeae (Pitter) Beyer & L.G. Lohmann, distribuído pela América Central, e (v) S. coriaceum Beyer & L.G. Lohmann, restrito ao Estado do Pará, na Amazônia Oriental. O estudo biogeográfico indicou que o ancestral de Stizophyllum e seu grupo-irmão Martinella, estava distribuído pela Amazônia. A divergência destas linhagens ocorreu durante o Eoceno, enquanto a diversificação das espécies de Stizophyllum ocorreu durante o Mioceno, a partir de um ancestral amplamente distribuído pela região Neotropical. Essa dissertação traz novos dados para um melhor entendimento dos processos que levaram à estruturação da biota Neotropical e contribui dados importantes para um projeto multidisciplinar amplo neste tópico (FAPESP 2012/50260-6) / The Neotropics is one of the most biodiverse regions in the planet. This region has a complex geological history that began during the break of Gondwana, which separated the South American and African continents in the Mesozoic, at ca. 150 million years ago. All the geological changes that followed greatly impacted the diversification of the fauna and flora of this region. However, it is still not clear what processes led to the high diversity found in this region. This study focuses on Stizophyllum (Bignonieae, Bignoniaceae), a small genus of Neotropical lianas, distributed from Mexico to southern Brazil. Although Stizophyllum is well circumscribed, species limits remain complicated in this group and little is known about its evolutionary history. This study aims to: (i) reconstruct the phylogeny of the genus and use it as a basis to infer the biogeographical history of this group, and (ii) develop nuclear and plastid microsatellite markers (SSRs) for future studies on the phylogeography and population genetics of this group. To this end, we reconstructed the phylogeny of the genus based on three molecular markers (ndhF, rpl32-trnL and pepC) and a broad sample of individuals, using Bayesian and Maximum Likelihood approaches. We then estimated divergence times of the various lineages and recontructed the biogeographical history of the group. Lastly, we developed nuclear microsatellite markers (nSSRs) and chloroplast microsatellite markers (cpSSRs) for the group. In total, we developed thirty-seven SSRs, nine from the nucleus and twenty-eight from the chloroplast. All markers amplified successfully in S. riparium and transferability was sucessful to S. inaequilaterum and S. perforatum. Five main clades were reconstructed in the molecular phylogeny of the group, all of which are characterized by good morphological markers and here recognized as species in an updated synopsis of the group: (i) S. inaequilaterum Bureau & K. Schum., distributed through the Amazon and Central America; (ii) S. perforatum (Cham.) Miers, distributed through the Atlantic Forest and the Dry Areas from Central Brasil; (iii) S. riparium (Kunth) Sandwith, distributed throughout the Amazon Basin; (iv) S. flos-ardeae (Pitter) Beyer & L.G. Lohmann, distributed through Central America; and, (v) S. coriaceum Beyer & L.G. Lohmann, restricted to the state of Pará, in Eastern Amazonia. The biogeographic study indicated that the ancestor of Stizophyllum and its sister-group Martinella was broadly distributed through Amazônia. These lineages dievrsified during the Eocene, while the diversification of Stizophyllum species occurred during the Miocene, from an ancestor that was broadly distributed through the Neotropics. This dissertation brings new information for the assembly of the Neotropical biota and contributes important data for a broader multidisciplinary project on this topic (FAPESP 2012 / 50260-6)

A região Neotropical é uma das regiões com maior biodiversidade no planeta. Essa região apresenta uma complexa história geológica que iniciou-se com a quebra da Gondwana, separando os continentes sul-americano e africano, durante o Mesozóico, há ca. de 150 milhões. Todas as mudanças geológicas que seguiram influenciaram a diversificação da fauna e flora dessa região. No entanto, ainda não sabemos quais processos levaram à alta diversidade encontrada nesta região. Esse estudo, foca em Stizophyllum, (Bignonieae, Bignoniaceae), um pequeno gênero de lianas Netropicais, que ocorre desde o México até o sul do Brasil. Apesar da cirscunscrição de Stizophyllum ser clara, limites específicos permanecem complicados neste grupo e pouco se sabe sobre sua história evolutiva. Este estudo visa: (i) reconstruir a filogenia do gênero e utilizá-la como base para inferir a história biogeográfica do grupo, e (ii) desenvolver marcadores de microssatélites (SSRs) nucleares e plastidiais, para futuros estudos filogeográficos e de genética de populações. Para tal, reconstruímos a filogenia do gênero com base em três marcadores moleculares (ndhF, rpl32-trn,L e pepC) e uma ampla amostragem de indivíduos, utilizando inferências bayesiana e de máxima verossimilhança. Em seguida estimamos as idades de divergência das diversas linhagens e reconstruímos a história biogeográfica do grupo. Por fim, desenvolvemos marcadores de microssatélite nucleares (nSSRs) e de cloroplasto (cpSSRs) para o grupo. Ao todo, desenvolvemos trinta e sete SSRs, nove nucleares e vinte oito de cloroplasto. Todos marcadores foram polimórficos em S. riparium e apresentaram sucesso de transferabilidade para S. inaequilaterum e S. perforatum. Cinco clados principais foram reconstruídos na filogenia molecular do gênero, os quais são caracterizados por bons caracteres morfológicos e aqui reconhecidos como espécies em uma nova sinopse apresentada para o grupo: (i) S. inaequilaterum Bureau & K. Schum., distribuído pela Amazônia e América Central; (ii) S. perforatum (Cham.) Miers, distribuído pela Mata Atlântica e Áreas Secas do Brasil Central, (iii) S. riparium (Kunth) Sandwith, distribuído por toda Bacia Amazônia; (iv) S. flos-ardeae (Pitter) Beyer & L.G. Lohmann, distribuído pela América Central, e (v) S. coriaceum Beyer & L.G. Lohmann, restrito ao Estado do Pará, na Amazônia Oriental. O estudo biogeográfico indicou que o ancestral de Stizophyllum e seu grupo-irmão Martinella, estava distribuído pela Amazônia. A divergência destas linhagens ocorreu durante o Eoceno, enquanto a diversificação das espécies de Stizophyllum ocorreu durante o Mioceno, a partir de um ancestral amplamente distribuído pela região Neotropical. Essa dissertação traz novos dados para um melhor entendimento dos processos que levaram à estruturação da biota Neotropical e contribui dados importantes para um projeto multidisciplinar amplo neste tópico (FAPESP 2012/50260-6) / The Neotropics is one of the most biodiverse regions in the planet. This region has a complex geological history that began during the break of Gondwana, which separated the South American and African continents in the Mesozoic, at ca. 150 million years ago. All the geological changes that followed greatly impacted the diversification of the fauna and flora of this region. However, it is still not clear what processes led to the high diversity found in this region. This study focuses on Stizophyllum (Bignonieae, Bignoniaceae), a small genus of Neotropical lianas, distributed from Mexico to southern Brazil. Although Stizophyllum is well circumscribed, species limits remain complicated in this group and little is known about its evolutionary history. This study aims to: (i) reconstruct the phylogeny of the genus and use it as a basis to infer the biogeographical history of this group, and (ii) develop nuclear and plastid microsatellite markers (SSRs) for future studies on the phylogeography and population genetics of this group. To this end, we reconstructed the phylogeny of the genus based on three molecular markers (ndhF, rpl32-trnL and pepC) and a broad sample of individuals, using Bayesian and Maximum Likelihood approaches. We then estimated divergence times of the various lineages and recontructed the biogeographical history of the group. Lastly, we developed nuclear microsatellite markers (nSSRs) and chloroplast microsatellite markers (cpSSRs) for the group. In total, we developed thirty-seven SSRs, nine from the nucleus and twenty-eight from the chloroplast. All markers amplified successfully in S. riparium and transferability was sucessful to S. inaequilaterum and S. perforatum. Five main clades were reconstructed in the molecular phylogeny of the group, all of which are characterized by good morphological markers and here recognized as species in an updated synopsis of the group: (i) S. inaequilaterum Bureau & K. Schum., distributed through the Amazon and Central America; (ii) S. perforatum (Cham.) Miers, distributed through the Atlantic Forest and the Dry Areas from Central Brasil; (iii) S. riparium (Kunth) Sandwith, distributed throughout the Amazon Basin; (iv) S. flos-ardeae (Pitter) Beyer & L.G. Lohmann, distributed through Central America; and, (v) S. coriaceum Beyer & L.G. Lohmann, restricted to the state of Pará, in Eastern Amazonia. The biogeographic study indicated that the ancestor of Stizophyllum and its sister-group Martinella was broadly distributed through Amazônia. These lineages dievrsified during the Eocene, while the diversification of Stizophyllum species occurred during the Miocene, from an ancestor that was broadly distributed through the Neotropics. This dissertation brings new information for the assembly of the Neotropical biota and contributes important data for a broader multidisciplinary project on this topic (FAPESP 2012 / 50260-6)

Recent theoretical developments have led to a renewed interest in the potential role of chromosomal rearrangements in speciation. Australian morabine grasshoppers (genus Vandiemenella, viatica species group) provide an excellent study system to test this potential role, because they show extensive chromosomal variation: 12 chromosomal races/species with parapatric distributions. The research in this thesis involves the application of molecular genetic analyses to examine patterns of gene introgression among chromosomal races of Vandiemenella at three different spatial scales: local-scale hybrid zone analysis, island-scale phylogeography, and continental-scale phylogeography. The aims of these multi-scale analyses are to investigate whether chromosomal races represent genetically distinct taxa with limited gene flow, and to infer the historical biogeography of Vandiemenella and evolutionary origins of their parapatric distributions. Karyotype and 11 nuclear markers revealed a remarkably narrow hybrid zone with substantial linkage disequilibrium and strong deficits of heterozygotes between the chromosome races P24(XY) and viatica17 on Kangaroo Island, suggesting that the zone is maintained by a balance between dispersal and selection against hybrids (tension zone). Selection that maintains the stable hybrid zone is unlikely to be operating only on loci linked to rearranged chromosomes. Island-scale and continental-scale phylogeography using multiple nuclear markers indicated that Vandiemenella chromosome races/species generally represent genetically distinct taxa with reduced gene flow between them. In contrast, analyses of a mitochondrial gene showed the presence of distinctive and geographically localised phylogroups that do not correspond with the distribution of the Vandiemenella taxa. These discordant population genetic patterns are likely to result from introgressive hybridization between the taxa and range expansions and contractions. Overall, our molecular analyses favour the allopatric mode of diversification for the evolution of Vandiemenella and do not support the stasipatric speciation model of White (1978). Patterns of genetic differentiation between the chromosomal races analysed at three different spatial scales show dynamic responses of the grasshoppers to past climatic fluctuations, leading to opportunities for long-term isolation and allopatric fixation of new chromosome variants and molecular mutations at many loci. Further analyses are necessary to assess potential roles of chromosomal rearrangements in facilitating diversification in Vandiemenella by reducing recombination within the rearranged chromosome segments.

Total antioxidant activity and total carotenoid levels were evaluated for more than 100 common potato (Solanum tuberosum, L.) cultivars grown in the United States, advanced breeding lines from several Western U.S. breeding programs, and 47 related, tuber-bearing species. An initial assessment of variability for antioxidant activity provided baseline information to be used for potential potato promotion and for the development of new varieties with greater human health benefits. Wide variability in antioxidant levels provided evidence of genetic control of this trait, indicating that it could be possible to breed for enhanced levels of antioxidant compounds in potato. Accessions, varieties, and advanced breeding lines identified in the broad screen as having high antioxidant activity and high total carotenoid levels, were fine screened via HPLC to determine specific phenolic and carotenoid compounds present in potato. The objective of the study was to identify parents for use in the Texas breeding program to develop potato varieties containing increased levels antioxidant compounds. In the broad screen for total antioxidant activity, the 47 related, tuber-bearing species showed a wider range of variability than the cultivated varieties and breeding lines. Based on the DPPH assay, antioxidant activity ranged from 103-648 uM trolox equivalents in the cultivated varieties and advanced breeding lines, while that of the wild species was 42-892. HPLC analysis revealed that the phenolic content of the species, and their cultivated counterparts, was primarily composed of caffeic and chlorogenic acids. Other phenolics identified were p-coumaric acid, rutin hydrate, vanillic acid, epicatechin, t-cinnamic acid, gallic acid, and salicylic acid. The highest phenolic content discovered in the accessions was five-fold higher than the highest of the cultivated genotypes. Carotenoid analysis revealed lutein in the accessions, but the yellow-flesh breeding lines were much higher in carotenoids. In addition to the work conducted on antioxidants, an attempt was made to separate intraclonal variants of the potato cultivar Russet Norkotah. Eleven microsatellite primers and 112 AFLP primer combinations failed to produce any reproducible polymorphisms. The inability to detect differences between the clones could be due to the tetraploid nature of the clones or epigenetic differences not detected by the procedures utilized in this study.

The domestic dog, Canis lupus familiaris, has emerged as a model system for the study of human hereditary diseases. Of the approximately 450 hereditary diseases described in the dog, half have clinical presentations that are quite similar to specific human diseases. Understanding the genetic bases of canine hereditary diseases will not only complement comparative genetics studies but also facilitate selective breeding practices to reduce incidences in the dog. Whole genome screens have great potential to identify the marker(s) that segregate with canine hereditary diseases for which no reasonable candidate genes exist. The Minimal Screening Set-1 (MSS-1) was the first set of microsatellite markers described for linkage analysis in the dog and was, until recently, the best tool for genome screens. The MSS-2 is the most recently described screening set and offers increased density and more polymorphic markers. The first objective of this work was to develop tools to streamline genomic analyses in the study of canine hereditary diseases. This was achieved through the development of 1) multiplexing strategies for the MSS-1, 2) a multiplex of microsatellite markers for use in canine forensics and parentage assays and 3) chromosome-specific multiplex panels for the MSS-2. Multiplexing is the simultaneous amplification and analysis of markers and significantly reduces the expense and time required to collect genotype information. Pancreatic acinar atrophy (PAA) is a disease characterized by the degeneration of acinar cells of the exocrine pancreas and is the most important cause of exocrine pancreatic insufficiency (EPI) in the German Shepherd Dog (GSD). Although the prognosis for dogs having EPI is typically good with treatment, many dogs are euthanized because the owners are unable to afford the expensive enzyme supplements. The second objective of this work was to determine the mode of transmission of EPI in the GSD and conduct a whole genome screen for linkage. Two extended families of GSDs having PAA were assembled and used to determine the pattern of transmission. The results of this indicate that PAA is an autosomal recessive disease. The multiplexed MSS-1 was used to conduct an initial whole genome screen, although no markers were suggestive of linkage.

The primary goal of the Thoroughbred industry is to breed and train superior equine athletes capable of excelling on the racetrack. To date, research into the genetic underpinnings of athletic ability has been limited in the horse. Advances in equine genomics and the genetics of athletic performance in humans have opened up the possibility of investigating this important trait in the Thoroughbred. Initially, 46 candidate genes associated with human athletic performance were mapped in the equine genome by radiation hybrid (RH) and fluorescent in situ hybridization (FISH) mapping. RH data and later the draft equine genomic sequence allowed us to identify microsatellites adjacent to these and other candidate genes (95 in total). Additional microsatellites were added to increase genome coverage, producing a final panel of 186 markers. All the potential markers were initially screened on a pool of DNA for 16 Thoroughbreds to ensure they were polymorphic. The panel was genotyped on 162 Thoroughbreds in total; Centimorgans (cM) between microsatellites were determined with CRI-MAP. The animal?s athletic ability was estimated using career winnings loge transformed to create a linear trait; unraced animals were treated as missing data. Linkage analysis was carried out using the MERLIN program, and association analysis was carried out using the QTDT program. Appropriate thresholds for statistical significance were determined by carrying out 1000 simulated genome scans based on the structure of the original data. LOD scores above 1.54 met the criteria of statistical significance (with a 5% chance of type I error). In the actual genome scan, the marker L12.2 had the highest observed LOD score of 1.16 and p-value of 0.01 and consequently was not significant; the association analysis also did not detect significant association with performance on the track. Given the complexity of the phenotype under investigation and the modest sample size, the lack of linkage/association was not unexpected. Nevertheless, this study has contributed to the RH and FISH maps of the equine genome. Additionally, the development of the genome scanning panel for this study has provided useful information on the most informative microsatellites for linkage or association studies in the Thoroughbred.

Total antioxidant activity and total carotenoid levels were evaluated for more than 100 common potato (Solanum tuberosum, L.) cultivars grown in the United States, advanced breeding lines from several Western U.S. breeding programs, and 47 related, tuber-bearing species. An initial assessment of variability for antioxidant activity provided baseline information to be used for potential potato promotion and for the development of new varieties with greater human health benefits. Wide variability in antioxidant levels provided evidence of genetic control of this trait, indicating that it could be possible to breed for enhanced levels of antioxidant compounds in potato. Accessions, varieties, and advanced breeding lines identified in the broad screen as having high antioxidant activity and high total carotenoid levels, were fine screened via HPLC to determine specific phenolic and carotenoid compounds present in potato. The objective of the study was to identify parents for use in the Texas breeding program to develop potato varieties containing increased levels antioxidant compounds. In the broad screen for total antioxidant activity, the 47 related, tuber-bearing species showed a wider range of variability than the cultivated varieties and breeding lines. Based on the DPPH assay, antioxidant activity ranged from 103-648 uM trolox equivalents in the cultivated varieties and advanced breeding lines, while that of the wild species was 42-892. HPLC analysis revealed that the phenolic content of the species, and their cultivated counterparts, was primarily composed of caffeic and chlorogenic acids. Other phenolics identified were p-coumaric acid, rutin hydrate, vanillic acid, epicatechin, t-cinnamic acid, gallic acid, and salicylic acid. The highest phenolic content discovered in the accessions was five-fold higher than the highest of the cultivated genotypes. Carotenoid analysis revealed lutein in the accessions, but the yellow-flesh breeding lines were much higher in carotenoids. In addition to the work conducted on antioxidants, an attempt was made to separate intraclonal variants of the potato cultivar Russet Norkotah. Eleven microsatellite primers and 112 AFLP primer combinations failed to produce any reproducible polymorphisms. The inability to detect differences between the clones could be due to the tetraploid nature of the clones or epigenetic differences not detected by the procedures utilized in this study.

Advances in molecular technologies have provided conservation biologist with the opportunity to quantify the genetic structure of a population and, in turn develop management guidelines and policies aimed at preserving the genetic diversity of fish stocks challenged by human activities. This thesis examines the status of genetics as applied to the management of freshwater fisheries by state natural resource agencies with a purpose of understanding the keys to a successful genetics program. An online survey was used to investigate the breadth of molecular marker application to freshwater fisheries management by state natural resource departments. Seven questions were posed to 50 state agencies addressing species of concern, type of genetic resources used, type of molecular marker used, and management concerns. Genetics was listed as a concern in the management of 18 freshwater fish families representing 70 distinct species, with Salmonid species the most frequently reported (20%#37;). A majority of agencies rely on outside resources to perform genetics testing (65%#37;). The most common analysis technique used by state agencies was microsatellite DNA analysis (35%#37;) and the most frequently reported management concerns were genetic stock identification and management boundaries (23%#37;). The application of a specific molecular technology to a conservation question was addressed by investigating the mechanisms of unnatural selection in the form of a study of trait heritability. Microsatellite parentage analysis was used to reconstruct familial relationships of juvenile Florida bass (Micropterus floridanus) displaying variable traits of growth and aggressiveness in a culture setting. Differences in the parentage of high growth and aggression (HGA) and baseline growth and aggression (BGA) offspring showed that certain parent-pairings contribute disproportionally to certain size classes and levels of aggression. These results suggest that the selective pressures of recreational harvest may negatively impact the fitness of wild fish stocks. Overall, this work provides natural resource managers with the basic information required to successfully develop and employ strategies aimed at preserving the genetic integrity of freshwater fisheries.

The critically endangered Kemp's ridley sea turtle experienced a dramatic decline in population size (demographic bottleneck) between 1947 and 1987 from 160,000 mature individuals to less than 5000. Demographic bottlenecks can cause genetic bottlenecks where significant losses of genetic diversity occur through genetic drift. The loss of genetic diversity can lower fitness through the random loss of adaptive alleles and through an increase in the expression of deleterious alleles. Molecular genetic studies on endangered species require collecting tissue using non-invasive or minimally invasive techniques. Such sampling techniques are well developed for birds and mammals, but not for sea turtles. The first objective was to explore the relative success of several minimally invasive tissue-sampling methods as source of DNA from Kemp's ridley sea turtles. Tissue sampling techniques included; blood, cheek swabs, cloacal swabs, carapace scrapings, and a minimally invasive tissue biopsy of the hind flipper. Single copy nuclear DNA loci were PCR amplified with turtle-specific primers. Blood tissue provided the best DNA extractions. Additionally, archival plasma samples are shown to be good sources of DNA. However, when dealing with hatchlings or very small individuals in field situations, the tissue biopsy of the hind flipper is the preferred method. This study's main focus was to evaluate whether the Kemp's ridley sea turtle sustained a measurable loss of genetic variation resulting from the demographic bottleneck. To achieve this goal, three alternative approaches were used to detect a reduction in Kemp's ridley's effective population size (Ne) from microsatellite data. These approaches were 1) Temporal change in allele frequencies, 2)An excess of heterozygotes in progeny, and 3)A mean ratio (M) of the number of alleles (k) to the range of allele size (r). DNA samples were obtained from Kemp's ridleys caught in the wild. PCR was used to amplify eight microsatellite loci and allele frequencies were determined. Data from only four microsatellites could be used. Although the reduced number of loci was a limiting factor in this study, the results of all three approaches suggest that Kemp's ridley sustained a measurable loss of genetic variation due to the demographic bottleneck.

The domestic dog, Canis lupus familiaris, has emerged as a model system for the study of human hereditary diseases. Of the approximately 450 hereditary diseases described in the dog, half have clinical presentations that are quite similar to specific human diseases. Understanding the genetic bases of canine hereditary diseases will not only complement comparative genetics studies but also facilitate selective breeding practices to reduce incidences in the dog. Whole genome screens have great potential to identify the marker(s) that segregate with canine hereditary diseases for which no reasonable candidate genes exist. The Minimal Screening Set-1 (MSS-1) was the first set of microsatellite markers described for linkage analysis in the dog and was, until recently, the best tool for genome screens. The MSS-2 is the most recently described screening set and offers increased density and more polymorphic markers. The first objective of this work was to develop tools to streamline genomic analyses in the study of canine hereditary diseases. This was achieved through the development of 1) multiplexing strategies for the MSS-1, 2) a multiplex of microsatellite markers for use in canine forensics and parentage assays and 3) chromosome-specific multiplex panels for the MSS-2. Multiplexing is the simultaneous amplification and analysis of markers and significantly reduces the expense and time required to collect genotype information. Pancreatic acinar atrophy (PAA) is a disease characterized by the degeneration of acinar cells of the exocrine pancreas and is the most important cause of exocrine pancreatic insufficiency (EPI) in the German Shepherd Dog (GSD). Although the prognosis for dogs having EPI is typically good with treatment, many dogs are euthanized because the owners are unable to afford the expensive enzyme supplements. The second objective of this work was to determine the mode of transmission of EPI in the GSD and conduct a whole genome screen for linkage. Two extended families of GSDs having PAA were assembled and used to determine the pattern of transmission. The results of this indicate that PAA is an autosomal recessive disease. The multiplexed MSS-1 was used to conduct an initial whole genome screen, although no markers were suggestive of linkage.