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Background Goat, one of the first domesticated livestock, is a worldwide important species both culturally and economically. The current goat reference genome, known as ARS1, is reported as the first nonhuman genome assembly using 69× PacBio sequencing. However, ARS1 suffers from incomplete X chromosome and highly fragmented Y chromosome scaffolds. Results Here, we present a very high-quality de novo genome assembly, Saanen_v1, from a male Saanen dairy goat, with the first goat Y chromosome scaffold based on 117× PacBio long-read sequencing and 118× Hi-C data. Saanen_v1 displays a high level of completeness thanks to the presence of centromeric and telomeric repeats at the proximal and distal ends of two-thirds of the autosomes, and a much reduced number of gaps (169 vs. 773). The completeness and accuracy of the Saanen_v1 genome assembly are also evidenced by more assembled sequences on the chromosomes (2.63 Gb for Saanen_v1 vs. 2.58 Gb for ARS1), a slightly increased mapping ratio for transcriptomic data, and more genes anchored to chromosomes. The eight putative large assembly errors (1 to ~ 7 Mb each) found in ARS1 were amended, and for the first time, the substitution rate of this ruminant Y chromosome was estimated. Furthermore, sequence improvement in Saanen_v1, compared with ARS1, enables us to assign the likely correct positions for 4.4% of the single nucleotide polymorphism (SNP) probes in the widely used GoatSNP50 chip. Conclusions The updated goat genome assembly including both sex chromosomes (X and Y) and the autosomes with high-resolution quality will serve as a valuable resource for goat genetic research and applications.

Die Anforderungen an molekulargenetische Untersuchungen sind in den letzten Jahren gewachsen, selbst aus schwierigem Proben- oder Spurenmaterial mit wenig und fragmentierter DNA sollen möglichst aufschlussreiche, genetische Informationen gewonnen werden. Y-chromosomal lokalisierte Single Nucleotide Polyorphisms (SNPs) können einen wichtigen Beitrag zur Erweiterung des forensischen Analysespektrums leisten, das bisher nahezu ausschließlich auf einem DNA-Mustervergleich von Spur und Referenzprobe basiert. In der vorliegenden Arbeit wird ein hierarchischer Analyseansatz vorgestellt, mit dem die durch Y-SNPs definierten Haplogruppen entlang der Phylogenie des Y-Chromosoms typisiert werden können und damit jede männliche DNA genau einem zeitlich und räumlich definierten Ast des genetischen Stammbaums des modernen Menschen zugeordnet werden kann. Dabei bietet die Kombination aus PCR- Multiplexanalysen und Minisequenzierung eine schnelle und effektive Untersuchungsmethode. Anhand von zwei Fallstudien, in denen Skelettfunde mit forensischem und archäogenetischem Hintergrund untersucht wurden, wird dargestellt, dass Y-SNPs sich besonders für Untersuchungen von degradiertem DNA-Material eignen. Es wird gezeigt, dass sich mit Y-SNPs aufgrund ihrer Populationsspezifität eine Vorhersage der geographischen Herkunft unbekannter DNA aus Spurenmaterial vornehmen lässt, und zwar besser als mit jedem anderen einzelnen genetischen Marker. Diese charakteristische Information aus dem Bereich der DNA intelligence kann bei Fällen ohne Vergleichsproben („cold cases“), sowie bei der Aufklärung von Vermisstenfällen zur sinnvollen Eingrenzung des Personenkreises verwendet werden. Um die geographische Verteilung der Haplogruppen zu ermitteln, ist die Durchführung von regionalen Populationsstudien wie sie hier für indigene und moderne Populationen durchgeführt wurden, unerlässlich. Dass diese Studien darüber hinaus auch zum Verständnis von Migration, Populationsgenese und prähistorischen sowie historischen Ereignissen in der Menschheitsgeschichte beitragen können, wird hier am Beispiel der Besiedlung Südamerikas demonstriert., A challenge of forensic casework is to meet the increasing demands on molecular analyses. Even low copy or strongly fragmented DNA should provide helpful genetic information for the purpose of solving criminal cases. The analysis of Y-linked Single Nucleotide Polymorphisms (SNPs) is a new approach to comply with most difficult cases. The method extends the current forensic DNA analysis spectrum by providing DNA intelligence information. In this work, a hierarchical assay to analyze SNP defined haplogroups along the phylogeny of the Y chromosome is presented, assigning each male DNA to a temporal and spatial specified branch of the genetic genealogy of modern humans. The combination of PCR-multiplexes and minisequencing provides an effective and sensitive approach. With the application of these techniques to two case studies, where skeletal remains were investigated with forensic and archaeogenetic background, it is shown that Y-SNPs are valuable genetic markers to analyze especially degraded DNA material. Furthermore, it could be demonstrated that due to the population specificity of Y-SNPs, the geographical origin of unknown male DNA can be identified much better than with any other single genetic marker. This “intelligence” information can support police investigations in “cold cases” in which no suspect is known and in missing person cases to narrow down the number of concerned persons. Population studies are essential for revealing the phylogeographical distribution of Y-chromosomal haplogroups. Here, authochthonous and admixed populations were examined for this purpose. The results of these studies also contribute to the understanding of the past demography of populations shaped by founder effects, migration, admixture and bottlenecks. This is illustrated by the example of the peopling of South America.

Short Tandem Repeats (STRs) sind DNA-Abschnitte, in denen sich eine kurze Nukleotid-Sequenz aufeinanderfolgend wiederholt. Das Genom des Menschen besteht zu etwa 3% aus solchen Sequenzen, und Mutationen an einigen dieser Loci sind für bestimmte Krankheiten verantwortlich. Wegen ihrer sehr hohen Mutationsraten werden STRs, insbesondere solche auf dem Y-Chromosom (Y-STRs), oft als genetische Marker verwendet, z.B. in der Forensik und für Vaterschaftstests. Viele dieser Anwendungen setzen Modelle für die STR-Evolution voraus. Solche Modelle bestehen aus der Kombination eines genealogischen Modells mit einem Mutationsmodell. Der für STRs wichtigste Mutationsmechanismus ist das Slipped-Strand Mispairing während der DNA-Replikation. Dies führt dazu, dass eine STR-Mutation in der Regel im Gewinn oder Verlust einer Repeat-Einheit besteht. Daraus ergibt sich als einfachstes STR-Mutationsmodell das Stepwise Mutation Model (SMM), welches das genealogische Wright-Fisher-Modell um einen entsprechenden Mutationsprozess erweitert. Mittels Markov-Ketten-Theorie charakterisieren wir das global wandernde, aber lokal kohärente Verhalten der Allelfrequenzen-Verteilung in einer Population, das unter dem SMM erwartet wird. Dazu wird der entsprechende Allelprozess X betrachtet, der sich als nullrekurrente Markov-Kette erweist (globales Verhalten). Eine modifizierte Version V dieses Prozesses, bei der von jedem Allel das Allel eines bestimmten Individuums (oder der Mittelwert aller Allele) der jeweiligen Generation subtrahiert wird, ist dagegen positiv rekurrent (lokales Verhalten). Die Markov-Kette V konvergiert exponentiell schnell gegen eine eindeutige unimodale stationäre Verteilung eta. Es wird gezeigt, wie man eta numerisch approximieren kann. Über das SMM hinaus existiert eine Vielzahl an STR-Mutationsmodellen. Viele davon berücksichtigen die Tatsache, dass die STR-Mutationsrate von der vorhandenen Allel-Länge abhängt. Bekannt ist aber auch, dass locus-spezifische Faktoren ebenfalls eine Rolle spielen. Um dies zu berücksichtigen, haben wir zu sechs Y-STR-Loci die Daten von 15.285 Vater-Sohn-Paaren aus der forensischen Literatur zusammengetragen. Dies ermöglicht die quasi direkte Beobachtung von 162 Mutationen. Mit einem Maximum-Likelihood-Ansatz werden anhand dieser Daten das SMM, das Lineare Modell (in dem die Mutationswahrscheinlichkeit linear mit der Allel-Länge des Vaters zunimmt) und ein neues Logistisches Modell in verschiedenen Versionen locus-spezifisch miteinander verglichen. Für fünf der sechs betrachteten Y-STRs zeigt sich, dass eine bestimmte Version unseres Logistischen Modells am besten passt. Es wird diskutiert, inwieweit diese Ergebnisse auf autosomale Loci übertragbar sind. Ein Beispiel für die praktische Anwendung von STR-Evolutionsmodellen ist das Schätzen der Random Match Probability (RMP) für die Quantifizierung der Evidenz einer Profilübereinstimmung im Rahmen forensischer DNA-Analysen. Dieses ist bei zuvor unbekannten Y-STR-Profilen (Singleton-Haplotypen) besonders problematisch. Für diesen Fall werden verschiedene RMP-Schätzer beschrieben und simulationsbasiert miteinander verglichen. Insbesondere betrachten wir eine koaleszenzbasierte Methode, die ein STR-Mutationsmodell voraussetzt. Dieser Schätzer ist zwar sehr rechenaufwändig, erweist sich den anderen Methoden ansonsten aber als überlegen. Abschließend wird diskutiert, ob STRs eine evolutionäre Funktion besitzen, bzw. wie groß der Anteil funktioneller Sequenzen am menschlichen Genom ist. Dies ist deshalb bedeutsam, weil z.B. in Deutschland nur neutrale Marker für Identifikationszwecke verwendet werden dürfen. Short tandem repeats (STRs) are stretches of DNA that consist of a tandemly repeated short nucleotide sequence. About 3% of the human genome are STRs, and mutations at some of these loci are responsible for certain diseases. Due to their exceptionally high mutation rates, STRs, in particular those on the Y chromosome (Y-STRs), are often used as genetic markers, e.g. in forensics and for paternity tests. Many of these applications require models of STR evolution. Such models are obtained by combining a genealogical model with a mutation model. Concerning STR mutation, slipped-strand mispairing during DNA replication is the most important mechanism. This results in STR mutations usually comprising the gain or loss of one repeat unit. The Stepwise Mutation Model (SMM), which augments the genealogical Wright-Fisher model accordingly, is thus the simplest STR mutation model. Applying Markov chain theory, we characterize the globally wandering but locally coherent behaviour of the allele frequency distribution in a population that is expected under the SMM. To this end, the appropriate allele process X is considered and shown to be a null recurrent Markov chain (global behaviour). On the other hand, a modified version V of this process, obtained by subtracting the allele of a specific individual (or the mean of all alleles) in the current generation from each allele, is positive recurrent (local behaviour). The Markov chain V converges exponentially fast towards a unique unimodal stationary distribution eta. It is shown how eta can be approximated numerically. Beyond the SMM, a multitude of STR mutation models exist. Many of these take into account the fact that the STR mutation rate depends on the given allele length. But it is also known that locus-specific factors play a role, too. To accomodate this, we accumulated data on six Y-STR loci in 15,285 father-son pairs from the forensics literature. These data allow the virtually direct observation of 162 mutations, and they are used for a locus-specific comparison of some STR mutation models via a maximum-likelihood approach. Considered are the SMM, the Linear Model (in which the mutation rate increases linearly with the father's allele length) and a novel Logistic Model in various versions. For five of the six Y-STRs, a certain version of our Logistic Model fits best. It is discussed in how far these results are generalizable to autosomal loci. One example for the practical use of STR evolution models is the estimation of the random match probability (RMP) to quantify the evidence of a profile match in forensic DNA typing. This estimation is especially problematic if the profile in question is a Y-STR haplotype that has not been observed previously (a singleton haplotype). Regarding this case, we describe several RMP estimators and conduct simulations to compare them. In particular, we consider a coalescent-based method that requires an STR mutation model. Although this estimator is computationally demanding, it performs best otherwise. Finally, the question whether STRs possess an evolutionary function is discussed. This question hinges on the proportion of functional sequences in the human genome. The importance of these considerations is underlined by the fact that in many countries, e.g. Germany, the law allows only neutral markers to be used for identification purposes.

In dieser Arbeit wurden 200 Skelette von einem frühchristlichen Kirchfriedhof aus dem 12. / frühen 13. Jahrhundert aus Usedom (Kr. Ostvorpommern, Land Mecklenburg-Vorpommern) untersucht. Die Stadt Usedom war im Belegungszeitraum ein wichtiger Seehandelsplatz und Ort großer politischer und sozialer Veränderungen. Die Annahme des Christentums durch die slawische Elite im Jahr 1128, die folgenden Überfälle der Dänen sowie der Beginn des Zuzugs fränkisch- deutscher Siedler ab dem 13. Jahrhundert waren prägende Ereignisse. Mit Hilfe gängiger anthropologischer und paläodemographischer Methoden erfolgte die Rekonstruktion der Lebensbedingungen der historischen Usedomer Bevölkerung. Die Alters- und Geschlechtsverteilung zeigen eine für die Zeit normale Bevölkerungsstruktur mit einem hohen Kinderanteil (32 %), verhältnismäßig wenigen Jugendlichen und vielen Erwachsenen (59 %) sowie einem leichten Männerüberschuss. Bemerkenswert ist eine mit einem Frauendefizit in der maturen Altersklasse einhergehende Übersterblichkeit der Mädchen der Altersklasse infans I, welcher allerdings auch ein methodischer Deutungsfehler zugrunde liegen kann. Um einer Klärung des ethnischen Hintergrundes der slawisch, dänisch und schließlich deutsch geprägten Bevölkerung näher zu kommen, wurden acht Schädelmaße, vier Schädelindizes und 5 Langknochenmaße sowohl typologisch als auch statistisch anhand ihres Mittelwertunterschieds mit den Maßen von zwei Vergleichsserien aus dem slawischen Sanzkow und der dänischen Seehandelstadt Haithabu verglichen, in der eine ethnisch stark gemischten Bevölkerung lebte. Der Mittelwertvergleich sowohl der Schädelmaße und –indizes als auch der Langknochenmaße sowie der Körperhöhe erbrachte einige signifikante Unterschiede zwischen den Knochenmaßen der Bewohner von Usedom, Sanzkow und Haithabu. Diese Unterschiede deuten auf eine größere Ähnlichkeit der Usedomer Frauen und Männer mit der slawischen Vergleichspopulation hin, wobei die Maße der Usedomer eine Mittelstellung zwischen den Vergleichsserien einzunehmen scheinen. Ein osteometrischer Vergleich mit Schädel- und Langknochenmaßen von slawischen und germanischen Populationen aus der Zeit vom Neolithikum bis in das frühe Mittelalter brachte keine tiefergehenden Ergebnisse. Der archäologisch fundierten Aussage, dass die Bevölkerung noch überwiegend slawisch geprägt sei, wird aus anthropologischer Sicht nicht widersprochen. Die Analyse der mitochondrialen DNA (mtDNA) und Y-chromosomalen DNA war trotz widriger Lagerungsbedingungen erfolgreich. Von allen vier untersuchten Usedomern konnte die Haplogruppe der mtDNA (2 x H, 2 x K) bestimmt werden. Die Y-chromosomale Analyse ergab die Haplogruppen E1b1b und R1a1a7. Die Resultate lassen bisher leider nur den Schluss zu, dass es sich bei den Usedomern um Europäer handelt. In naher Zukunft wird die molekulargenetische Forschung mit genaueren Analysemethoden die Migrationsbewegungen der Menschen durch die Jahrhunderte immer besser rekonstruieren können., This study investigates 200 skeletons from an early Christian graveyard of the 12th to early 13th century in Usedom (Mecklenburg-Vorpommern, Germany). The city of Usedom was a notable maritime place of trade in a time of major political and social transformations. The Christianisation of the Slavic elite in 1128, the following raids of the Danes and the influx of German settlers starting in the 13th century were formative events. The reconstruction of the living conditions of the Usedom population was achieved by means of well established anthropological and palaeodemographical methods. Age and sex distribution comply with other ordinary populations of that time frame: high proportion of children (32 %), comparatively few adolescents but many adults (59 %) as well as a slight surplus in men. Remarkably, a deficit in women in the mature age class is attended by an increased mortality of girls of the age class infans I. However, this may be due to a methodical error. In order to clarify a possible Slavic, Danish or German background of the inhabitants of Usedom, eight skull measures, four skull indices and five measures of the long bones of the extremities were investigated typologically as well as statistically on the basis of their arithmetic means and compared to the measures of two series of Slavic or multiethnic/place of trade background (Sanzkow and Haithabu, respectively). The comparison of arithmetic means did yield statistically significant differences between the three populations. The men and women of Usedom seem to be more closely related to the Sanzkow population. However, they appear to take a position between the two other populations. Unfortunately, a comparison with Slavic and Germanic populations of the Neolithic till Early Middle Ages did not provide distinct results. The archaeologically based assumption of a mainly Slavic population cannot be rejected with anthropological means. The analysis of mitochondrial and Y-chromosomal DNA, however, generated auspicious results despite adverse storage conditions. Results could be obtained from all four samples. Two individuals were of mtDNA haplogroup H and two of haplogroup K. Y-chromosome analysis yielded haplogroups E1b1b and R1a1a7, respectively, in two males. Future molecular research will see improved methods for the even more detailed reconstruction of human migration.

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The authors describe 3 cases where Y-chromosomal systems were used for typing the biological traces. In the first case, a murder, for the major amount of cell material found on a dish towel (and analysed two years after the crime) female persons were excluded for the system amelogenin and Y-chromosomal systems. A brother of the victim could not be excluded for autosomal STR-systems. Upon confrontation with the results of the DNA-analysis (among other things), this brother confessed the murder of his sister some days later. He was found guilty by the court. In the second case described, a rape of two girls, many traces were analysed parallely with Y-chromosomal and autosomal PCR-systems. The objects where male DNA matching the suspect were found (a paper tissue, a sweat shirt and the knickers of the girls), also showed small amounts of alleles matching with the suspect for autosomal systems, while the major part in these systems was from the girls. The suspect was sentenced to many years imprisonment. In the third case, a possible rape of a young woman, a stained microscope slide of a vaginal swab had to be examined. Microscopically a few sperm heads could be seen in a surplus of leucocytes. The male proportion could be analysed only in the Y-chromosomal systems, not in the autosomal ones. For the frequency calculation of the Y-chromosomal allele combination the haplotype data bank of the Institute for Legal Medicine of the Humboldt university in Berlin was indispensable.

Myelodysplastic syndromes (MDS) are a heterogenous group of clonal stem cell disorders which generally occur in older adults but may also affect children. Primary MDS should be distinguished from secondary MDS associated with antineoplastic or immunosuppressive therapy (t-MDS), exposure to toxic compounds, or genetic disorders. The establishment of a neoplastic clone is reflected by dysplastic features and impaired function which may affect all three hematopoietic cell lineages. The ineffective hematopoiesis which causes bone marrow failure is accompanied by peripheral blood cytopenia and is considered to result from increased apoptosis, at least in the less advanced MDS stages. The elucidation of the molecular pathogenesis of MDS has provided evidence that chromosomal abnormalities are present in about 50% of patients with primary MDS. They include numerical aberrations such as monosomy 5 or 7, trisomy 8, loss of the Y-chromosome and structural abnormalities such as deletion of the long arm of chromosome 5 (5q-syndrome), 7, or 8. Based on the percentage of blasts (5%, 5-20%, 20-30%) and the presence of15% ringed sideroblasts for marrows with5% blasts, the French-American-British (FAB) classifies MDS into 4 morphologic categories: refractory anemia (RA), refractory anemia with excess of blasts (RAEB), refractory anemia with excess of blasts in transformation (RAEB-t), and refractory anemia with ringed sideroblasts. The fifth morphologic type is chronic myelomonocytic leukemia characterized by peripheral blood monocytosis (1x10(9)/l). However, a modification of this classification will be proposed by the World Health Organization, with the intention of lowering the threshold for the diagnosis of AML from 30% to 20% blast cells. In patients presenting with cytopenias suggesting impaired hematopoiesis, the initial diagnosis depends mainly on the cytological evaluation of bone marrow and blood smears and the histological findings of trephine bone marrow biopsy. In a retrospective analysis we evaluated the occurrence of the distinct FAB-categories as percentage of the total number of MDS cases diagnosed at the Institute of Pathology of the University of Freiburg. A total of 63% fullfilled the criteria of RA/RARS, 17% of RAEB, 14% of RAEB-t, and 6% of CMML. A fibrotic variant of MDS was observed in 7.67% of all cases, ranging from 2.34% in RA up to 15. 42-15.84% in the categories which did not show significant differences with regard to myelofibrosis. The histologic evaluation of a trephine bone marrow biopsy is of critical importance for the evaluation of fibrotic or hypocellular MDS since these patterns are not reflected by the cytological examination. The combined cytological and histological diagnosis of bone marrow and peripheral blood is a reliable tool for the initial diagnosis of MDS. In addition, cytogenetic and molecular analysis should be performed. Presently, the risk of leukemic transformation is evaluated using the International Prognostic Scoring System for MDS, which is the sum of the scores of bone marrow blasts, karyotypes and cytopenia. In the context of clinical trials therapeutic modalities should be considerd according to the age and the general performance state and the prognostic scores of individual patients.

Renal cell tumors display notoriously an intra- and intertumoral morphological heterogeneity making a reproducible cytomorphological diagnosis impossible. To overcome the cytological heterogeneity, the "Heidelberg Classification of Renal Cell Tumours" is soundly based on the genetic lesions that underlie the formation of distinct tumors. Allelic loss of chromosome 3 p and mutation of the VHL gene specify conventional renal cell carcinoma. In addition duplication of chromosome 5 q as well as allelic losses of prognostic value at chromosomes 6 q, 8 p, 9 p and 14 q characterize conventional renal cell carcinomas. Papillary renal cell tumors, which display an adenoma-carcinoma sequence, are marked by trisomies of chromosomes 3 q, 7, 8 p, 12 q, 16 q, 17 q und 20 as well as loss of the Y chromosome in males. Chromophobe renal cell carcinoma shows a low chromosome number due to monosomy of chromosomes 1, 2, 6, 10, 13, 17, and 21. Benign tumors, such as renal oncocytoma and metanephric adenoma, can also be diagnosed by genetic means. The distinct types of tumors have different biological behaviour and therefore, an accurate classification is considered clinically important. We worked out a quick and reproducible fluorescent microsatellite assay for detecting allelic changes. This technique is useful for postoperative and retrospective diagnosis as well as for preoperative analysis of fine needle aspiration specimens.

If hermaphrodite genitals are present in the patient or a higher degree of hypospadia is shown with maldescensus testis, a chromosomal disorder must be considered as one potential cause of the anomaly. The case report of a child with cryptorchidism on the right, inguinal testis on the left and penoscrotal hypospadia is presented as an example. A mosaic karyotype 45, X/46, X, idic (Yp) was diagnosed in this patient after chromosomal analysis. The cell line with the isodicentric Y chromosome could be demonstrated in about 90% of the lymphocytes, but only in 7% of the fibroblasts of the preputium. A derivative Y could not be detected in interphase nuclei in the buccal mucosa, i.e. only the cell line with monosomy X was presented. There was thus chromosomal mosaicism with unequal tissue involvement and a high potential for malignant transformation. Guidelines of pediatric urological, cytogenetic and endocrinological investigations and the diagnostic procedures are described and discussed. A prevention protocol for patients with comparable gonosomal mosaicism is presented.

Chronic rejection is a major problem in contemporary kidney transplantation. The purpose of this study was to determine whether renal cells are repopulated by extra-renal cells over time or whether the graft remains permanently allogenic. We studied nine explanted allografted kidneys of sex-mismatched donors by means of non-isotopic in situ hybridization (NISH). We used biotinylated centromer-specific DNA probes of the human chromosomes Y and X. In a further step, monoclonal and polyclonal antibodies against CD45, CD3, CD20, CD31, CD1a, S100, alpha-actin, factor Vill and UEA were used to analyse the various infiltrating cell types and the cells involved in allograft arteriopathy. In several cases NISH and immunohistochemistry were combined to facilitate the typing of cells. Our study showed that up to several years after transplantation the glomerular, tubular and endothelial cells retained donor origin. The only cells of recipient origin were the inflammatory cells, predominantly macrophages and T lymphocytes.

Recent molecular studies on transitional cell carcinomas (TCC) of the urinary bladder suggest two genetically different tumor types with a varying degree of genomic instability-pTa (grade 1 and 2) tumors are usually "genetically stable". These tumors have rarely p53 alterations and contain only few genomic alterations (deletions, DNA sequence copy number gains, amplifications). Genomic changes in these tumors typically include losses of the chromosomes Y and 9 as well as 1q gains. TCC's with invasive growth pattern (pT1-4) are mostly "genetically instable". They have p53 alterations in about 50% of tumors, and often numerous DNA sequence copy number changes including deletions of 2q, 5q, 6q, 8p and 11p as well as gains at 1q, 3p, 3q, 5p, 6p, 8q und 10p. High level amplifications at various sites of the genome are also frequent in these "instable tumors". They may pinpoint the localization of overexpressed oncogenes. Further studies will have to investigate whether genetic analyses can add useful information to the currently used pathological (pTNM) and clinical (superficial/invasive) classifications of urinary bladder cancer.

In order to elucidate cytogenetic changes associated with the development of benign growth of follicular epithelial cells of the thyroid, cytogenetic analyses were performed on 340 follicular thyroid adenomas and goiters after short-term culture. Clonal chromosome changes were found in 67 cases. Trisomy 7 as the sole abnormality or along with other trisomies was the most frequent type of aberration (19 cases). Other recurrent numerical changes were loss of chromosome 22 (4 cases) and the second X or the Y chromosome (5 cases). Translocations involving 19q13 (12 cases) were frequent structural chromosome changes. After a histopathological classification of all cases, we have correlated the cytogenetic findings with the histology of the tumors. Only 8.4% of the goiters showed clonal abnormalities whereas 44.9% of the adenomas revealed clonal abnormalities. Furthermore, simple clonal changes were predominantly found in goiters and complex changes in adenomas. The most impressive correlation was found in the group of lesions with trisomy 7: Whereas all but one lesion with one or two additional trisomies were goiters, those having three or more additional trisomies were all adenomas or adenomatous goiters.

Genetically directed representational difference analysis (GDRDA) allows to isolate DNA fragments that differ between complex genomes. This approach was applied in the present work to isolate Y-specific bovine sequences. Of eight isolated and cloned fragments, two proved to be Y-specific.

The percentage of freemartins among blood samples tested by chromosome analysis amounted to 83.9%, by blood group serologie 71.4%. 126 blood samples have been tested by blood group serology and PCR. Employing blood group serology, 71.3% and using PCR with BOV97M primers 85.8% of the animals proved to be freemartins. 40 blood samples were additionally analysed using PCR with zinc-finger-gene primers. 36 animals (90%) were identified as being freemartins by means of BOV97M and 34 animals (85%) by means of the zinc-finger-gene primer. The PCR method proved to be a rapid and very sensitive method for the diagnosis of freemartins and also suitable for routine testing. The BOV97M primer showed to have a higher Y chromosome specificity than the zinc-finger-gene primer.

aDNA-analyses are presented for ancient bones, teeth, soft tissues and dried body fluids. The implications and perspectives for Historical Anthropology are discussed. Methods are described on examples of molecular sex determination and kinship analysis. Finally, first amplifications of DNA-sequences from pathogenes of tuberculosis and plague from ancient skeletal material are reported.

The genetics of human sex determination is considered in view of the various disorders of gonad development. The Y chromosome plays an important role in the induction of sex determination by encoding the testis-determining factor (TDF). However, not all deviations in regular development can be explained by mutations of the TDF as unique factor. Therefore, it is necessary to postulate other mutations in still unknown genes of the cascade for male-specific determination as well as the requirement of an ovary-determining factor for regular female development.

This is a case report on a 19-year-old patient with the chromosome constellation 48, XYYY. The patient showed signs of dysmorphism, mental retardation and problems in psychosocial functioning. A literature review revealed that individuals with this karyotype have widely varying phenotypes with regard not only to physical signs but also to brain functions and cognitive performance.

We report on three family members suffering from both autosomal dominant Hailey-Hailey disease and bipolar affective disorder. As molecular biology techniques have made the localization of genes causing simple Mendelian traits possible as a routine task, the gene for Hailey-Hailey disease will presumably be localized in the foreseeable future. The Hailey-Hailey gene and its chromosomal surrounding will then be a region of interest for linkage studies in bipolar affective disorder.

With spontaneous abortion the conceptus is often expelled and lost right at the beginning, and uterine curettings then contain only endometrial fragments and clotted blood. Even complete embedding of all available material for histologic examination will not reveal any chorionic villi, and ectopic pregnancy can thus not be excluded. In such cases, the intermediate trophoblast can sometimes still be demonstrated within the endometrial tissue. This highly invasive trophoblast is difficult to identify using conventional staining, but cytokeratin antibodies are reliable markers of this cell type. Using immunohistochemistry, these fetal components could be demonstrated in 27 of 95 specimens (28.5%), proving the intrauterine nature of the aborted pregnancy. In some cases the fetal derivation of intermediate trophoblast was demonstrated by using in situ hybridisation to mark repetitive sequences on the Y-chromosome in the interphase nucleus.

In bone marrow transplantation (BMT) the detection of residual host lymphoid or haematopoietic cells surviving conditioning therapy is because of its association to graft-versus-host disease, graft-versus-leukemia reaction, and relapse of leukemia a matter of great interest. We studied the occurrence of this mixed lymphoid chimerism (MC) in the formol-fixed lymphatic tissue of lymph nodes and spleen from 21 autopsies after allogeneic sex-mismatched BMT (5 females, 16 males, survival 5 to 1140 days after BMT). In situ hybridisation with biotinylated centromer-specific anti-X- and anti-Y-chromosome probes was performed on pepsin-digested paraffin sections. The number of double X-, single X-, and Y-chromosome bearing cells was analysed microscopically. Because of artefacts only 14 cases remained for valid investigation. MC was detected in 6 cases (5 out of 11 males 5 days to 840 days and 1 out of 3 females 76 days after BMT). MC occurred after whole body irradiation with 10 Gy (n = 5) and 7 Gy (n = 1). In 1 autopsy relapse of leukemia caused host cell infiltration. Cases with MC did not express histological signs of acute or chronic graft-versus-host disease, but 5 out of 8 with complete lymphoid chimerism did. The sensitivity of interphase cytogenetics on paraffin embedded tissue is low.

Thirty-one routinely processed radical prostatectomy specimens were examined for the presence of numerical chromosomal aberrations by in situ hybridization with centromeric nucleic acid probes specific for chromosomes 7, 10, 17, X, and Y. In eight of the prostatectomy specimens, chromosome numbers consistent with a normal male karyotype were found. Three cases, in addition to diploid chromosome numbers, showed a focal doubling of hybridization signals, consistent with tetraploidy. The other 20 cases displayed more or less marked numerical chromosomal aberrations. In this group, the appearance of numerical chromosomal aberrations often showed considerable local heterogeneity and was significantly correlated with tumor stage, Gleason grades, and tumor volume. We conclude that in prostatic cancer the presence of numerical chromosomal aberrations is associated with advanced disease. Especially in low differentiated tumors local heterogeneity in chromosome numbers can be very marked.

We report on a 14 year-old boy with 48,XXYY karyotype, presenting with essential tremor and a slight retardation of psychomotor development. In contrast to other cases with 48,XXYY syndrome, this pubertal patient has testicles of normal size, has small stature and shows no behavioural disturbances. We compared our patient with 120 other published cases.

Separation of X- and Y-chromosome-bearing spermatozoa has been attempted using Sephadex gel filtration. Fractions rich in X-chromosome-bearing spermatozoa were eluted in dependence on length of separation distance and on speed of elution. Spermatozoa remaining at the gel showed a higher percentage of Y-bodies compared with the untreated control. Sedimentation pattern and amount of eluted cells is influenced by eluation conditions.