A gene-based high-resolution comparative radiation hybrid map as a framework for genome sequence assembly of a bovine chromosome 6 region associated with QTL for growth, body composition, and milk performance traits

BACKGROUND: A number of different quantitative trait loci (QTL) for various phenotypic traits, including milk production, functional, and conformation traits in dairy cattle as well as growth and body composition traits in meat cattle, have been mapped consistently in the middle region of bovine chromosome 6 (BTA6). Dense genetic and physical maps and, ultimately, a fully annotated genome sequence as well as their mutual connections are required to efficiently identify genes and gene variants responsible for genetic variation of phenotypic traits. A comprehensive high-resolution gene-rich map linking densely spaced bovine markers and genes to the annotated human genome sequence is required as a framework to facilitate this approach for the region on BTA6 carrying the QTL. RESULTS: Therefore, we constructed a high-resolution radiation hybrid (RH) map for the QTL containing chromosomal region of BTA6. This new RH map with a total of 234 loci including 115 genes and ESTs displays a substantial increase in loci density compared to existing physical BTA6 maps. Screening the available bovine genome sequence resources, a total of 73 loci could be assigned to sequence contigs, which were already identified as specific for BTA6. For 43 loci, corresponding sequence contigs, which were not yet placed on the bovine genome assembly, were identified. In addition, the improved potential of this high-resolution RH map for BTA6 with respect to comparative mapping was demonstrated. Mapping a large number of genes on BTA6 and cross-referencing them with map locations in corresponding syntenic multi-species chromosome segments (human, mouse, rat, dog, chicken) achieved a refined accurate alignment of conserved segments and evolutionary breakpoints across the species included. CONCLUSION: The gene-anchored high-resolution RH map (1 locus/300 kb) for the targeted region of BTA6 presented here will provide a valuable platform to guide high-quality assembling and annotation of the currently existing bovine genome sequence draft to establish the final architecture of BTA6. Hence, a sequence-based map will provide a key resource to facilitate prospective continued efforts for the selection and validation of relevant positional and functional candidates underlying QTL for milk production and growth-related traits mapped on BTA6 and on similar chromosomal regions from evolutionary closely related species like sheep and goat. Furthermore, the high-resolution sequence-referenced BTA6 map will enable precise identification of multi-species conserved chromosome segments and evolutionary breakpoints in mammalian phylogenetic studies

The synergistic interaction of thermal stress coupled with overstocking strongly modulates the transcriptomic activity and immune capacity of rainbow trout (Oncorhynchus mykiss)

The objective of the present study is to identify and evaluate informative indicators for the welfare of rainbow trout exposed to (A) a water temperature of 27 °C and (B) a stocking density of 100 kg/m3 combined with a temperature of 27 °C. The spleen-somatic and condition index, haematocrit and the concentrations of haemoglobin, plasma cortisol and glucose revealed non-significant differences between the two stress groups and the reference group 8 days after the onset of the experiments. The transcript abundance of almost 1,500 genes was modulated at least twofold in in the spleen of rainbow trout exposed to a critical temperature alone or a critical temperature combined with crowding as compared to the reference fish. The number of differentially expressed genes was four times higher in trout that were simultaneously challenged with high temperature and crowding, compared to trout challenged with high temperature alone. Based on these sets of differentially expressed genes, we identified unique and common tissue- and stress type-specific pathways. Furthermore, our subsequent immunologic analyses revealed reduced bactericidal and inflammatory activity and a significantly altered blood-cell composition in challenged versus non-challenged rainbow trout. Altogether, our data demonstrate that heat and overstocking exert synergistic effects on the rainbow trout’s physiology, especially on the immune system.publishedVersio

Analysis of Autozygosity Using Whole-Genome Sequence Data of Full-Sib Families in Pikeperch (Sander lucioperca)

Pikeperch (Sander lucioperca) has emerged as a high value species to the aquaculture industry. However, its farming techniques are at an early stage and its production is often performed without a selective breeding program, potentially leading to high levels of inbreeding. In this study, we identified and characterized autozygosity based on genome-wide runs of homozygosity (ROH) on a sample of parental and offspring individuals, determined effective population size (N ( e )), and assessed relatedness among parental individuals. A mean of 2,235 ± 526 and 1,841 ± 363 ROH segments per individual, resulting in a mean inbreeding coefficient of 0.33 ± 0.06 and 0.25 ± 0.06 were estimated for the progeny and parents, respectively. N ( e ) was about 12 until four generations ago and at most 106 for 63 generations in the past, with varying genetic relatedness amongst the parents. This study shows the importance of genomic information when family relationships are unknown and the need of selective breeding programs for reproductive management decisions in the aquaculture industry

Application of disease-associated differentially expressed genes – Mining for functional candidate genes for mastitis resistance in cattle

In this study the mRNA differential display method was applied to identify mastitis-associated expressed DNA sequences based on different expression patterns in mammary gland samples of non-infected and infected udder quarters of a cow. In total, 704 different cDNA bands were displayed in both udder samples. Five hundred-and-thirty two bands, (75.6%) were differentially displayed. Ninety prominent cDNA bands were isolated, re-amplified, cloned and sequenced resulting in 87 different sequences. Amongst the 19 expressed sequence tags showing a similarity with previously described genes, the majority of these sequences exhibited homology to protein kinase encoding genes (26.3%), to genes involved in the regulation of gene expression (26.3%), to growth and differentiation factor encoding genes (21.0%) and to immune response or inflammation marker encoding genes (21.0%). These sequences were shown to have mastitis-associated expression in the udder samples of animals with and without clinical mastitis by quantitative RT-PCR. They were mapped physically using a bovine-hamster somatic cell hybrid panel and a 5000 rad bovine whole genome radiation hybrid panel. According to their localization in QTL regions based on an established integrated marker/gene-map and their disease-associated expression, four genes (AHCY, PRKDC, HNRPU, OSTF1) were suggested as potentially involved in mastitis defense

Cloning and expression of activation induced cytidine deaminase from Bos taurus'

Activation induced cytidine deaminase is an enzyme crucial to somatic hypermutation and gene conversion, processes that are essential for the diversification of Ig V genes. The bovine Ig repertoire appears to be diversified by mechanisms that are significantly different to those that operate in humans and mice. This study set out to test the hypothesis that differences in the organization, coding sequence, expression or genomic location of the bovine AICDA gene enables the encoded enzyme to catalyse the unusual Ig diversification mechanism seen in cattle as well as conventional antigen-driven mutation. Characterization of bovine AICDA excluded the first two possibilities. AICDA expression was detected in lymphoid tissues from neonatal and older cattle, but AICDA cDNA could not be detected in muscle tissue. The pattern of gene expression did not therefore differ from that in other vertebrates. The AICDA cDNA was cloned and expressed successfully in Escherichia coli generating a phenotype consistent with the mutating action of this deaminase. Using a whole genome radiation hybrid panel, bovine AICDA was mapped to a region of bovine chromosome 5 syntenic with the location of human AICDA on chromosome 12. We conclude that the unusual nature of Ig diversification in cattle is unlikely to be attributable to the structure, sequence, activity or genomic location of bovine AICDA

A connectome and analysis of the adult Drosophila central brain.

The neural circuits responsible for animal behavior remain largely unknown. We summarize new methods and present the circuitry of a large fraction of the brain of the fruit fly Drosophila melanogaster. Improved methods include new procedures to prepare, image, align, segment, find synapses in, and proofread such large data sets. We define cell types, refine computational compartments, and provide an exhaustive atlas of cell examples and types, many of them novel. We provide detailed circuits consisting of neurons and their chemical synapses for most of the central brain. We make the data public and simplify access, reducing the effort needed to answer circuit questions, and provide procedures linking the neurons defined by our analysis with genetic reagents. Biologically, we examine distributions of connection strengths, neural motifs on different scales, electrical consequences of compartmentalization, and evidence that maximizing packing density is an important criterion in the evolution of the fly's brain

Under control: The innate immunity of fish from the inhibitors' perspective

The innate immune response involves a concerted network of induced gene products, preformed immune effectors, biochemical signalling cascades and specialised cells. However, the multifaceted activation of these defensive measures can derail or overshoot and, if left unchecked, overwhelm the host. A plenty of regulatory devices therefore mediate the fragile equilibrium between pathogen defence and pathophysiological manifestations. Over the past decade in particular, an almost complete set of teleostean sequences orthologous to mammalian immunoregulatory factors has been identified in various fish species, which prove the remarkable conservation of innate immune-control concepts among vertebrates. This review will present the current knowledge on more than 50 teleostean regulatory factors (plus additional fish-specific paralogs) that are of paramount importance for controlling the clotting cascade, the complement system, pattern-recognition pathways and cytokine-signalling networks. A special focus lies on those immunoregulatory features that have emerged as potential biomarker genes in transcriptome-wide research studies. Moreover, we report on the latest progress in elucidating control elements that act directly with immune-gene-encoding nucleic acids, such as transcription factors, hormone receptors and micro- and long noncoding RNAs. Investigations into the function of teleostean inhibitory factors are still mainly based on gene-expression profiling or overexpression studies. However, in support of structural and in-vitro analyses, evidence from in-vivo trials is also available and revealed many biochemical details on piscine immune regulation. The presence of multiple gene copies in fish adds a degree of complexity, as it is so far hardly understood if they might play distinct roles during inflammation. The present review addresses this and other open questions that should be tackled by fish immunologists in future

Xenogeneic and Stem Cell-Based Therapy for Cardiovascular Diseases: Genetic Engineering of Porcine Cells and Their Applications in Heart Regeneration

Cardiovascular diseases represent a major health concern worldwide with few therapy options for ischemic injuries due to the limited regeneration potential of affected cardiomyocytes. Innovative cell replacement approaches could facilitate efficient regenerative therapy. However, despite extensive attempts to expand primary human cells in vitro, present technological limitations and the lack of human donors have so far prevented their broad clinical use. Cell xenotransplantation might provide an ethically acceptable unlimited source for cell replacement therapies and bridge the gap between waiting recipients and available donors. Pigs are considered the most suitable candidates as a source for xenogeneic cells and tissues due to their anatomical and physiological similarities with humans. The potential of porcine cells in the field of stem cell-based therapy and regenerative medicine is under intensive investigation. This review outlines the current progress and highlights the most promising approaches in xenogeneic cell therapy with a focus on the cardiovascular system

Novel transcripts discovered by mining genomic DNA from defined regions of bovine chromosome 6

BACKGROUND: Linkage analyses strongly suggest a number of QTL for production, health and conformation traits in the middle part of bovine chromosome 6 (BTA6). The identification of the molecular background underlying the genetic variation at the QTL and subsequent functional studies require a well-annotated gene sequence map of the critical QTL intervals. To complete the sequence map of the defined subchromosomal regions on BTA6 poorly covered with comparative gene information, we focused on targeted isolation of transcribed sequences from bovine bacterial artificial chromosome (BAC) clones mapped to the QTL intervals. RESULTS: Using the method of exon trapping, 92 unique exon trapping sequences (ETS) were discovered in a chromosomal region of poor gene coverage. Sequence identity to the current NCBI sequence assembly for BTA6 was detected for 91% of unique ETS. Comparative sequence similarity search revealed that 11% of the isolated ETS displayed high similarity to genomic sequences located on the syntenic chromosomes of the human and mouse reference genome assemblies. Nearly a third of the ETS identified similar equivalent sequences in genomic sequence scaffolds from the alternative Celera-based sequence assembly of the human genome. Screening gene, EST, and protein databases detected 17% of ETS with identity to known transcribed sequences. Expression analysis of a subset of the ETS showed that most ETS (84%) displayed a distinctive expression pattern in a multi-tissue panel of a lactating cow verifying their existence in the bovine transcriptome. CONCLUSION: The results of our study demonstrate that the exon trapping method based on region-specific BAC clones is very useful for targeted screening for novel transcripts located within a defined chromosomal region being deficiently endowed with annotated gene information. The majority of identified ETS represents unknown noncoding sequences in intergenic regions on BTA6 displaying a distinctive tissue-specific expression profile. However, their definite regulatory function has to be analyzed in further studies. The novel transcripts will add new sequence information to annotate a complete bovine genome sequence assembly, contribute to establish a detailed transcription map for targeted BTA6 regions and will also be helpful to dissect of the molecular and regulatory background of the QTL detected on BTA6