Hybridization is an important mechanism in diversification. It often makes taxonomy difficult. Lack of strong supported intrageneric classification in genus Pteris (Pteridaceae) could be caused by natural hybridization. Most hybridization documnted in Pteris was based on limited evidence. This study focuses on Pteris cadieri complex, the taxon with putative hybridization. The species complex displayed significant morphological variation and was associated with hybrid origin. Reproductive biology revealed variation in spore number per sporangium, spore size, spore shape and apogamous reproduction, which imply its hybrid origin. Cytology analysis using chromosome counting and flow cytometry identified diploids, triploids, and tetraploids. CpDNA and nuclear DNA supported that Pteris cadieri complex is hybrid origin: paternal and maternal lineages were inferred and 11 taxa were identified. Furthermore, comparing materials form Hainan and Taiwan, , it is clear that the species complex is composed by taxa arisen from multiple hybridization. Systematic inconsistency existed between chloroplast and nuclear phylogenies in Pteris impled that other taxa might have involved in hybridization events, in addition to the Pteris cadieri complex. Hybridization may be very common in Pteris. To infer intrageneric taxonomy of Pteris, effect of reticulate evolution should never be neglected. Finally, based on morphological and evolutionary traits, the taxonomy of Pteris cadieri complex is revised. There are Pteris cadieri Christ, Pteris dimorpha Copel. var. dimorpha, Pteris dimorpha var. plumbea (Christ) Y.-S. Chao, H.-Y. Liu & W.-L. Chiou, Pteris grevilleana Wall. ex Agardh var. grevilleanan, Pteris grevilleana Wall. ex Agardh var. ornata Alderw., and Pteris hainanensis Ching.

The study of the DNA has found application beyond our understanding of its cellular function and into a variety of materials assembly and nucleic acid detection systems. The current research investigates double-stranded DNA probes in both a colloidal particle assembly and fluorescent assay format utilizing competitive hybridization events. In both contexts, the affinity of the dsProbes is tuned by the sequence design parameters of duplex length and complementarity. These systems were incubated with nucleic acid targets of interest and, based on the mechanism of competitive hybridization, were responsive to the presence of a high affinity competitive target. In the case of the particle assemblies, incubation with the competitive target resulted in observable disassembly of particle structures. In the case of fluorescently labeled dsProbes, incubation with competitive targets resulted in a quantifiable loss of fluorescence as determined by flow cytometry. Utilizing the fluorescently labeled dsProbe system, the kinetics of competitive hybridization was characterized for nucleic acid targets of varying specificity and strand context. The results indicate promise for the development of the competitive hybridization approach in nucleic acid detection systems providing advantages over current single-stranded probe designs. By utilizing a fluorescently labeled dsProbe approach, it is unnecessary to chemically modify the target of interest to impart a signaling mechanism. Additionally, as the process of competitive hybridization of dsProbes with targets of interest is an affinity driven process, discrimination of targets based on specificity is decoupled from standard measures such as elevated temperature protocols, an important step in translating nucleic acid technologies from the controlled laboratory environment to field applications.

The Aristolochiaceae are one of the largest angiosperm families, the family has been divided into two subfamilies: Asaroideae, which include Asarum and Saruma, and Aristolochioideae, which includes Thottea sensu lato and Aristolochia sensu lato (Kelly and Gonzales, 2003). Aristolochia sensu lato comprise between 450 and 600 species, distributed throughout the world with centers of diversities in the tropical and subtropical regions (Neinhuis et al., 2005, Wanke et al., 2006a, 2007). However, the extended Mediterranean region including Turkey, the Caucasus and the Near East is likely to be the only diversity hotspot of the genus Aristolochia in the northern hemisphere were up to 60 species and subspecies could be observed (Wanke 2007). Most important contributions to the knowledge of these species were published by Nardi (1984, 1988, 1991, 1993) and Davis & Khan (1961, 1964, 1982), all of these studies were based on morphological characters only. In recent years, with the progress of molecular techniques and in light of the systematic chaos, a detailed study was needed to unravel the evolutionary history prior to a taxonomic revision of this group. The first chapter of my thesis should be regarded as the starting point for more detailed investigation on population level. Preliminary molecular phylogenitic analysis recovered the Mediterranean Aristolochia species as monophyletic (de Groot et al 2006). However, only very few members were included in that study. The latest phylogenetic study by Wanke (2007) dealed with west Mediterranean Aristolochia species and sampled also few members belonging to the east Mediterranean and Caucasian species (3 from Greece, 2 from Georgia and 1 from Turkey). This study reported the Mediterranean Aristolochia species as two molecular and morphologically well supported clades, which were sister to each other. Furthermore, the two closely related species A. sempervirens and A. baetica which have an east west vicariance and are known as Aristolochia sempervirens complex has been recovered as sister group to the remaining west Mediterranean species. A detailed investigation of the evolutionary history of this group is the topic of the second chapter of my thesis (Chapter 2). The Aristolochia sempervirens complex is characterized by an unusual growth form and has a circum Mediterranean distribution. The investigation of these species complex seem to be of great importance to understand speciation and colonization of the Mediterranean by the genus Aristolochia and might shade light in historical evolutionary processes of other plant lineages in the Mediterranean. Furthermore, I test applicability and phylogenetic power of a nuclear single copy gene (nSCG) region to reconstruct well resolved and highly supported gene genealogies as a prerequisite to study evolutionary biology questions in general. Furthermore, a comprehensive overview of leaf epicuticular waxes, hairs and trichomes of 54 species from the old and new world taxa of the genus Aristolochia were investigated using scanning electron microscopy (SEM) to clarify taxonomic status of theses species in contrast to their molecular position. Also this study which is the third chapter of this thesis (Chapter 3), has a strong focus on Mediterranean Aristolochia and tries to provide additional support for molecular findings based on epicuticular waxes and to test them as synapomorphies. Each chapter has its own introduction and abstract resulting in a short general introduction here.

Mutations in the presenilin 1 gene account for most early-onset familial Alzheimer's disease (FAD). The presenilins and AD may also be related through a common involvement in the cell cycle. Here we report that one important aspect of the cell cycle---proper chromosome segregation---is dependent on presenilin function and therefore may be involved in AD pathogenesis. Specifically we find that FAD mutations in presenilin 1 (M146L and M146V) lead to chromosome missegregation and aneuploidy in vivo and in vitro: 1) Both metaphase chromosome analysis and in situ hybridization reveal significant aneuploidy in the lymphocytes and neurons of PS-1 transgenic mice. 2) Transiently transfected human cells expressing normal and, especially, mutant PS-1 develop aneuploidy within 48 hours, including trisomy 21, while cells transfected with dominant negative PS-1 genes lacking ?-secretase activity have no effect on chromosome segregation. 3) Analysis of mitotic spindles in the transfected cells reveals abnormal microtubule arrays and lagging chromosomes. The possible mechanisms by which cell cycle defects and chromosome missegregation induced by y-secretase may contribute to Alzheimer's disease will be discussed.

Circulating tumor cells (CTCs) are known to escape from the primary tumor site and may settle down at the distant organ to grow a second tumor. CTCs are one of causes initiating carcinoma metastasis. Detection of CTCs has been considered to be valuable for cancer management, including diagnosis, prognosis, and clinical treatment management. However, efficient isolation, enumeration, characterization, and genetic analysis of CTCs in whole-blood samples from cancer patients are very challenging due to their extremely low concentration and rare nature (per CTC in blood cells is 1:106–109). With the increasing worldwide death rate associated with cancer, there is a desperate demand for a high-sensitivity, high-throughput, and low-cost detection and separation system. My doctoral research focused on the design and fabrications of the screening system for the detection of CTCs with further analysis of captured CTCs, such as immunofluoresce staining and fluorescence in-situ hybridization (FISH). The distinct significance of this research is that the development of the computer-controlled rotational holder with a series of six inverted microfluidic chips reduced the cost by significantly reducing the consumption of magnetic carriers (25% of the consumed amount used in the commercial CellSearch® system), increasing the capture efficiency by manipulating the blood sedimentation in the microchannel, enhancing the system stability by integrating the micromagnets on the plain glass slide substrate, and achieving high throughput because of the high flow rate (2.5 mL/hr) and large screening volume (screening up to six chips in parallel with each containing 2.5 mL of blood). Immunofluorescence staining and the FISH method have been performed to prove the capability of the system. In addition, the system has been successfully applied for patient samples screening. The incorporation of micromagnets has demonstrated that micromagnets provide localized magnetic forces to scatter the target cancer cells and free nanoparticles throughout the whole channel substrate to increase the channel space usage by 13%. Four cancer cell lines, including COLO 205 (colorectal cancer), SK-BR-3 (breast cancer), MCF-7 (breast cancer), and PC3 (prostate cancer), were spiked in blood samples from healthy donors to verify high capture efficiency of the developed system. On average, over a 97% capture rate was demonstrated for all cell lines. Moreover, the developed screening system has been successfully screened over 40 patient samples, including metastatic lung cancer, breast cancer, prostate cancer, and colorectal cancer. After capture of CTCs, immunofluorescence staining was used to identified the captured cancer cells and the FISH method was performed to characterize the isolated cancer cells by studying the gene expression of CTCs from breast cancer. The proposed automated immunomagnetic microchip-based screening system shows high capture efficiency (average 97% for three spiked cell lines), high throughput (15 mL of blood sample per screening), high sensitivity, high specificity, and low nanoparticle consumption (75% less than CellSearch® system). The screening system provides great promise as a clinical tool for early cancer diagnosis, diagnosis, personalized therapy, and treatment monitoring. / text

<p>What evolutionary processes and ecological patterns underlie the maintenance of asexual reproduction in natural populations? Although a vast body of literature offers theory to explain the existence of sexual and asexual reproduction, there has been little study of these forms of reproduction in the natural environment. In this dissertation I use a combination of field experiments, greenhouse studies, and genetic techniques to answer this question in the model plant system Boechera. </p><p>In Chapter 1, I review the utility of this system for studying ecological and evolutionary questions in general. Boechera offers an array of genetic and genomic tools, facilitated in part by a close evolutionary relationship with the model plant Arabidopsis thaliana, as well as undisturbed ecology and habitat that extends across much of North America. Additionally, the presence of apomixis (asexual reproduction via parthenogenetic formation of seeds) at the diploid level makes Boechera an ideal system for studying sex without the often-confounding factor of polyploidy; Boechera is one of very few plant groups in which this is possible. </p><p>In Chapter 2, I use a combination of microsatellite markers, flow cytometry, chromosome squashes, and morphological work to characterize apomixis, polyploidy, and species diversity in over 100 natural populations collected from central Idaho and western Montana. As in many other apomictic systems, I find that apomixis in Boechera is strongly linked to hybridization between species or between genetically divergent intraspecific lineages. I then explore associations between apomixis and ecological and topographical variables, as well as variables underlying differentiation between apomictic and sexual lineages. I find that ecological variables associated with apomixis are largely in congruence with the hypothesis of geographic parthenogenesis, and that geographic parthenogenesis is likely driven by the consequences of interspecific hybridization. We also find that apomixis is linked with disturbance and slope, with apomicts occurring in flatter locations than sexuals. </p><p>In Chapter 3, I use a large-scale field experiment comprising three years of data from two cohorts of sexual and asexual lineages to compare fitness between these two groups. I find that, despite herbivory levels that are much higher in apomicts than sexuals, apomictic fitness is consistently higher than sexual. Viability selection strongly favors apomicts, which results in a total fitness advantage for apomicts, despite variable fecundity selection. Selection varies in intensity between cohorts and among gardens. The results of a complementary greenhouse experiment show that the effects of herbivory differ by reproductive mode. Together, these experiments suggest that Red Queen dynamics may contribute to the coexistence of sex and asex in this group.</p><p>In Chapter 4, I use inter- and intraspecific F2 crosses to conduct a greenhouse study and a field experiment to explore the effects of hybridization and heterozygosity on fitness. I find that heterozygosity is favored in the field, with viability selection strongly favoring outcrossed over inbred lineages. However, hybridization results in lower survival, reproduction, and total fitness of interspecific F2 crosses, although hybrids that do reproduce produce more fruits than selfed parental lineages of both species, resulting in fecundity selection for hybrids. It is clear that the benefits of apomixis are due to hybridization, as hybrids are less fit overall. Evidence for both heterosis and outbreeding depression, dependent on lineage and on trait, are found in the field; these phenomena are not clearly associated with geographic distance between parental populations. Ongoing SNP genotyping will facilitate assessment of heterozygosity-fitness correlations as well as correlation of fitness and heterozygosity.</p> / Dissertation

This dissertation presents the applications of state-of-the-art computation techniques and data analysis algorithms in three physical and biological problems: assembling DNA pieces, optimizing self-assembly yield, and identifying correlations from large multivariate datasets. In the first topic, in-depth analysis of using Sequencing by Hybridization (SBH) to reconstruct target DNA sequences shows that a modified reconstruction algorithm can overcome the theoretical boundary without the need for different types of biochemical assays and is robust to error. In the second topic, consistent with theoretical predictions, simulations using Graphics Processing Unit (GPU) demonstrate how controlling the short-ranged interactions between particles and controlling the concentrations optimize the self-assembly yield of a desired structure, and nonequilibrium behavior when optimizing concentrations is also unveiled by leveraging the computation capacity of GPUs. In the last topic, a methodology to incorporate existing categorization information into the search process to efficiently reconstruct the optimal true correlation matrix for multivariate datasets is introduced. Simulations on both synthetic and real financial datasets show that the algorithm is able to detect signals below the Random Matrix Theory (RMT) threshold. These three problems are representatives of using massive computation techniques and data analysis algorithms to tackle optimization problems, and outperform theoretical boundary when incorporating prior information into the computation. / Engineering and Applied Sciences

Die Evolution der heutigen globalen Diversität wurde in den letzten Millionen Jahren insbesondere durch klimatische Schwankungen und entsprechende Veränderungen biologischer Lebensräume beeinflusst. Besonders deutlich ist der Einfluss von Glazialen und Interglazialen auf die Evolution von Organismen der Nördlichen Hemisphäre. Weniger klar hingegen ist, inwiefern sich diese klimatischen Verhältnisse auf (sub-) tropische Regionen ausgewirkt haben, insbesondere auf das Afrikanische Savannen Biom. Unabhängig davon, haben Umweltveränderungen in tropischen und nicht-tropischen Gebieten zur Entstehung vieler der gegenwärtigen Hybridzonen geführt, in denen zuvor geographisch separierte Populationen in sekundären Kontakt gekommen sind. Obwohl Hybridisierung im Tierreich inzwischen nicht mehr als ein rares Phänomen betrachtet wird, ist das tatsächliche Ausmaß und die Bedeutung von Hybridisierung in der Evolution von Tieren noch lange nicht vollständig geklärt. Die Verbreitung Grüner Meerkatzen der Gattung Chlorocebus reflektiert nahezu die Ausdehnung Afrikanischer Savannengebiete und basierend auf Beobachtungen im Freiland sowie auf morphologischen Merkmalen von Museumsexemplaren, hybridisieren die meisten der sechs anerkannten parapatrischen Arten in ihren jeweiligen Kontaktzonen. Aufgrund dieser Eigenschaften habe ich in meiner Doktorarbeit Grüne Meerkatzen als Modellsystem genutzt, um zum einen die Bedeutung von Hybridisierung in der Evolution von Primaten und Tieren im Allgemeinen zu untersuchen, und zum anderen, um wesentliche Trends in der Evolution von Savannensäugetieren zu analysieren. Um in einem ersten Schritt grundlegende Informationen über die mitochondriale Diversität und Verbreitung der Grünen Meerkatzen zu erlangen, generierte ich vollständige Cytochrom b Sequenzen von Proben, die alle sechs Arten der Gattung und große Teile der gesamten Verbreitung Grüner Meerkatzen repräsentierten. Des Weiteren nutzte ich Sequenz-Informationen zweier Y-chromosomaler Loci, ein Fragment der sex determining region (SRY) und das letzte Intron des Zinc finger (ZFY), um eventuell zeitlich zurückliegende Hybridisierungsereignisse nachzuweisen. Um räumliche sowie zeitliche phylogeographische Muster zu rekonstruieren, habe ich basierend auf den bisher gewonnen Daten weitere mitochondriale Marker von selektiven Proben aller mitochondrialer Kladen sequenziert. Abschließend habe ich die Phylogeographie Grüner Meerkatzen mit den Phylogeographien dreier anderer weit verbreiteter Savannensäugetier-Gattungen verglichen, mit Pavianen (Papio), Warzenschweinen (Phacochoerus), und Kuhantilopen (Alcelaphus). Meine Analysen der mitochondrialen Daten lassen neun klar abgegrenzte Kladen erkennen, die keiner bisher vorgeschlagenen Taxonomie entsprechen. Zahlreiche para- und polyphyletische Beziehungen, verursacht durch nicht übereinstimmende Verbreitungsmuster mitochondrialer Kladen und morphologischer Merkmale, liefern Hinweise auf anhaltende introgressive Hybridisierung in den Kontaktzonen aller Arten, mit Ausnahme der beiden westafrikanischen Arten. Darüber hinaus weisen die Ergebnisse der mitochondrialen Analysen auf potentiell vergangene introgressive Hybridisierungsereignisse hin, die geographisch nicht in Gebiete gegenwärtiger Kontaktzonen fallen. Dies kann im Falle von C. pygerythrus in Ostafrika anhand der Y-chromosomalen Daten bestätigt werden. Männchen basierte Introgression und nuclear swamping haben hier offensichtlich zu dem zytonukleären Aussterben eines historischen Taxons geführt. Aber nicht alle diskordante Muster in der mitochondrialen Phylogenie sind Anzeichen für zurückliegende introgressive Hybridisierung. Innerhalb der Verbreitung von C. tantalus weisen sowohl mitochondriale als auch Y-chromsomale Daten auf zwei klar separierte und morphologisch kryptische Taxa hin (ein westliches und ein östliches Taxon), ein Befund der möglicherweise auf das Vorhandensein einer neuen Grünen Meerkatzenart hindeutet. In Übereinstimmung mit mitochondrialen Ergebnissen weisen Y-chromosomale Daten ebenfalls keine Anzeichen für Hybridisierung zwischen C. sabaeus und der westlichen C. tantalus-Form in Westafrika auf. Entgegen früherer Annahmen stellt der Volta Fluss und seine nördlicheren Zuflüsse offenbar auf gesamter Länge eine geographische Barriere dar und nicht wie vorher angenommen nur im südlicheren Teil des Flussverlaufs. Die Phylogeographie Grüner Meerkatzen weist somit auf eine komplexe evolutionäre Geschichte hin. Aufgrund der phylogenetischen Rekonstruktionen und Datierungen gehe ich von einem Westafrikanischen Ursprung der Gattung vor ca. 2,46 Millionen Jahren aus. Des Weiteren liefert die Phylogenie Hinweise auf eine erst später folgende Ausbreitung bis nach Südafrika und auf zwei zeitlich getrennte Besiedelungen nordöstlicher Regionen, eine vom Westen und eine von eher südlicheren Regionen aus. Im Vergleich zu den anderen Savannensäugetieren können keine zeitlichen Übereinstimmungen in Aufspaltungsmustern gefunden werden, weder im Vergleich zwischen den Primaten noch zwischen den Ungulaten. Zudem fallen Aufspaltungen innerhalb der Gattungen zeitlich sowohl mit kalt-ariden als auch mit warm-humiden Perioden zusammen. Veränderungen in Populationsgrößen innerhalb der letzten 500.000 Jahre zeigen unterschiedliche Muster zwischen Primaten und Ungulaten, die wahrscheinlich mit verschiedenen ökologischen Anpassungen und Habitat Präferenzen zu erklären sind. Da außerdem keine klaren Zusammenhänge zwischen dem zeitlichen Auftreten von Populationsschwankungen und dem letzten Interglazialem bzw. Glazialen Maximum gefunden werden können, scheint ein Einfluss von regional geprägtem Klima in Afrika wahrscheinlicher. Zusammengefasst hat die Evolution Grüner Meerkatzen im frühen Pleistozän begonnen und klimatische Schwankungen im Quartär haben vermutlich zu wiederkehrenden Veränderungen in der Ausbreitung geführt. Diese begünstigte die Entstehung von sekundären Kontaktzonen und führte zu weit verbreiteter introgressiver Hybridisierung innerhalb der Gattung. Das zytonukleäre Aussterben ehemaliger Taxa oder Populationen als Ergebnis lang anhaltender introgressiver Hybridisierung und darauffolgendem nuclear swamping machen zudem den potentiellen Einfluss von Hybridisierung in der Evolution von Primaten und Tieren im Allgemeinen deutlich. Die Phylogeographien der Savannensäugetiere zeigen, dass Aufspaltungen innerhalb der Gattungen sehr wahrscheinlich durch ausgeprägt humide wie auch aride Bedingungen begünstigt wurden, die eher durch regionale Klimatische Veränderungen in Afrika zu erklären sind als durch Glaziale Zyklen und Klimaschwankungen der Nördlichen Hemisphäre. Die Ergebnisse meiner Doktorarbeit betonen die Notwendigkeit der Analyse von sowohl mütterlich als auch väterlich vererbten Markern in phylogeographischen Studien, um ein möglichst vollständiges Bild von evolutionären Prozessen, Hybridisierung, sowie von genetischer und taxonomischer Vielfalt zu erlangen.

Fortschritte in der molekulargenetischen Charakterisierung von Tumorerkrankungen haben in den letzten Jahren die klinische Praxis zunehmend beeinflusst. Das Ziel dieser Arbeit war die Untersuchung von astrozytären Tumoren der WHO-Grade II bis IV und ihre Subtypisierung anhand der gefundenen chromosomalen Aberrationen. Ferner sollte der Einfluss der gefundenen Aberrationen auf klinische Parameter wie das Gesamtüberleben oder die rezidivfreie Zeit untersucht werden. Dazu wurden paraffinfixierte Proben von insgesamt 184 primären astrozytären Tumoren (28 low-grade Astrozytome, 6 low-grade Oligoastrozytome, 50 anaplastische Astrozytome, 4 anaplastische Oligoastrozytome, 96 Glioblastoma multiforme) mit der Comparativen Genomischen Hybridisierung (CGH) untersucht. Häufige Aberrationen in allen Malignitätsgruppen stellten chromosomale Zugewinne auf Chromosom 7 sowie Verluste von Chromosom 10 und 9p dar. High-grade Astrozytome zeigten ferner häufig Zugewinne von Chromosom 19 und 20 sowie Verluste von 13q, 14q und 15q. WHO-Grad-II-Astrozytome wiesen häufig Zugewinne auf Chromosom 8 sowie Verluste von Chromosom 4q und 6q auf. Eine kürzeres Gesamtüberleben zeigten high-grade Gliome mit Verlusten von Chromosom 10q und Zugewinnen auf 7p. In Glioblastomen verursachten zusätzlich Zugewinne auf 7q sowie Verluste von 14q, in anaplastischen Astrozytomen zusätzlich Verluste von 10p ein verringertes Gesamtüberleben. WHO-Grad-II-Astrozytome zeigten bei Verlusten von 3p ein schlechteres Gesamtüberleben. Chromosomale Aberrationen, die zu einem verlängerten Gesamtüberleben führten, waren Verluste von 1p und Zugewinne von 10p in WHO-Grad-III-Tumoren. Die rezidivfreie Zeit wurde in high-grade Gliomen durch Zugewinne auf 7p und Verluste von 10p verringert. Eine Verkürzung der rezidivfreien Zeit in Glioblastomen zeigten außerdem Tumoren mit Verlusten von 7q, 10q und 14q. In anaplastischen Astrozytomen führten Verluste von 1p und 19q sowie Zugewinne auf 8q und 10p, in WHO-Grad-II-Astrozytomen Verluste von Chromosom 6 zu einer verlängerten Zeit bis zum Rezidiv. Die Anfertigung onkogenetischer Baummodelle stellte verschiedene genetische Wege der Tumorgenese dar. Ein Cluster war gekennzeichnet durch einen Verlust von 6q, ein weiterer wurde initialisiert durch den Verlust von 13q, der dritte durch den Verlust von 9p. Der vierte Cluster wurde charakterisiert durch Zugewinne auf Chromosom 7 und Verluste von Chromosom 10, während der fünfte Cluster Zugewinne auf 8q sowie Verluste von 4q aufwies.

The methods presented in this study are aimed at the identification of subpopulations (clones) of genetically similar cells within tissue samples through measurement of loci-specific Fluorescence in-situ hybridization (FISH) spot signals for each nucleus and analyzing cell spatial distributions by way of Voronoi tessellation and Delaunay triangulation to robustly define cell neighbourhoods. The motivation for the system is to examine lung cancer patient for subpopulations of Non-Small Cell Lung Cancer (NSCLC) cells with biologically meaningful gene copy-number profiles: patterns of genetic alterations statistically associated with resistance to cis-platinum/vinorelbine doublet chemotherapy treatment. Current technologies for gene-copy number profiling rely on large amount of cellular material, which is not always available and suffers from limited sensitivity to only the most dominant clone in often heterogeneous samples. Thus, through the use of FISH, the detection of gene copy-numbers is possible in unprocessed tissues, allowing identification of specific tumour clones with biologically relevant patterns of genetic aberrations. The tissue-wide characterization of multiplexed loci-specific FISH signals, described herein, is achieved through a fully automated, multicolour fluorescence imaging microscope and object segmentation algorithms to identify cell nuclei and FISH spots within. Related tumour clones are identified through analysis of robustly defined cell neighbourhoods and cell-to-cell connections for regions of cells with homogenous and highly interconnected FISH spot signal characteristics. This study presents experiments which demonstrate the system’s ability to accurately quantify FISH spot signals in various tumour tissues and in up to 5 colours simultaneously or more through multiple rounds of FISH staining. Furthermore, the system’s FISH-based cell classification performance is evaluated at a sensitivity of 84% and specificity 81% and clonal identification algorithm results are determined to be comparable to clone delineation by a human-observer. Additionally, guidelines and procedures to perform anticipated, routine analysis experiments are established.