Evaluating the impact of scoring parameters on the structure of intra-specific genetic variation using RawGeno, an R package for automating AFLP scoring

Background  

Since the transfer and application of modern sequencing technologies to the analysis of amplified fragment-length polymorphisms (AFLP), evolutionary biologists have included an increasing number of samples and markers in their studies. Although justified in this context, the use of automated scoring procedures may result in technical biases that weaken the power and reliability of further analyses.     

Detection of mitochondrial insertions in the nucleus (NuMts) of Pleistocene and modern muskoxen

Background  

Nuclear insertions of mitochondrial sequences (NuMts) have been identified in a wide variety of organisms. Trafficking of genetic material from the mitochondria to the nucleus has occurred frequently during mammalian evolution and can lead to the production of a large pool of sequences with varying degrees of homology to organellar mitochondrial DNA (mtDNA) sequences. This presents both opportunities and challenges for forensics, population genetics, evolutionary genetics, conservation biology and the study of DNA from ancient samples. Here we present a case in which difficulties in ascertaining the organellar mtDNA sequence from modern samples hindered their comparison to ancient DNA sequences.     

SNP HiTLink: a high-throughput linkage analysis system employing dense SNP data

Background  

During this recent decade, microarray-based single nucleotide polymorphism (SNP) data are becoming more widely used as markers for linkage analysis in the identification of loci for disease-associated genes. Although microarray-based SNP analyses have markedly reduced genotyping time and cost compared with microsatellite-based analyses, applying these enormous data to linkage analysis programs is a time-consuming step, thus, necessitating a high-throughput platform.     

Utility of EST-derived SSR in cultivated peanut (Arachis hypogaea L.) and Arachis wild species

Background  

Lack of sufficient molecular markers hinders current genetic research in peanuts (Arachis hypogaea L.). It is necessary to develop more molecular markers for potential use in peanut genetic research. With the development of peanut EST projects, a vast amount of available EST sequence data has been generated. These data offered an opportunity to identify SSR in ESTs by data mining.     

A knowledge discovery object model API for Java

Background  

Biological data resources have become heterogeneous and derive from multiple sources. This introduces challenges in the management and utilization of this data in software development. Although efforts are underway to create a standard format for the transmission and storage of biological data, this objective has yet to be fully realized.     

Global Epigenetic Changes Induced by SWI2/SNF2 Inhibitors Characterize Neomycin-Resistant Mammalian Cells

Background

Previously, we showed that aminoglycoside phosphotransferases catalyze the formation of a specific inhibitor of the SWI2/SNF2 proteins. Aminoglycoside phosphotransferases, for example neomycin-resistant genes, are used extensively as selection markers in mammalian transfections as well as in transgenic studies. However, introduction of the neomycin-resistant gene is fraught with variability in gene expression. We hypothesized that the introduction of neomycin-resistant genes into mammalian cells results in inactivation of SWI2/SNF2 proteins thereby leading to global epigenetic changes.

Methodology

Using fluorescence spectroscopy we have shown that the inhibitor, known as Active DNA-dependent ATPase A Domain inhibitor (ADAADi), binds to the SWI2/SNF2 proteins in the absence as well as presence of ATP and DNA. This binding occurs via a specific region known as Motif Ia leading to a conformational change in the SWI2/SNF2 proteins that precludes ATP hydrolysis. ADAADi is produced from a plethora of aminoglycosides including G418 and Streptomycin, two commonly used antibiotics in mammalian cell cultures. Mammalian cells are sensitive to ADAADi; however, cells stably transfected with neomycin-resistant genes are refractory to ADAADi. In resistant cells, endogenous SWI2/SNF2 proteins are inactivated which results in altered histone modifications. Microarray data shows that the changes in the epigenome are reflected in altered gene expression. The microarray data was validated using real-time PCR. Finally, we show that the epigenetic changes are quantized.

Significance

The use of neomycin-resistant genes revolutionized mammalian transfections even though questions linger about efficacy. In this study, we have demonstrated that selection of neomycin-resistant cells results in survival of only those cells that have undergone epigenetic changes, and therefore, data obtained using these resistant genes as selection markers need to be cautiously evaluated.     

CpGcluster: a distance-based algorithm for CpG-island detection

Background  

Despite their involvement in the regulation of gene expression and their importance as genomic markers for promoter prediction, no objective standard exists for defining CpG islands (CGIs), since all current approaches rely on a large parameter space formed by the thresholds of length, CpG fraction and G+C content.     

Showing their true colors: a practical approach to volume rendering from serial sections

Background  

In comparison to more modern imaging methods, conventional light microscopy still offers a range of substantial advantages with regard to contrast options, accessible specimen size, and resolution. Currently, tomographic image data in particular is most commonly visualized in three dimensions using volume rendering. To date, this method has only very rarely been applied to image stacks taken from serial sections, whereas surface rendering is still the most prevalent method for presenting such data sets three-dimensionally. The aim of this study was to develop standard protocols for volume rendering of image stacks of serial sections, while retaining the benefits of light microscopy such as resolution and color information.     

SAT,a flexible and optimized Web application for SSR marker development

Background  

Simple Sequence Repeats (SSRs), or microsatellites, are among the most powerful genetic markers known. A common method for the development of SSR markers is the construction of genomic DNA libraries enriched for SSR sequences, followed by DNA sequencing. However, designing optimal SSR markers from bulk sequence data is a laborious and time-consuming process.     

Validation of housekeeping genes for quantitative real-time PCR in in-vivo and in-vitro models of cerebral ischaemia

Background  

Studies of gene expression in experimental cerebral ischaemia models can contribute to understanding the pathophysiology of brain ischaemia and to identifying prognostic markers and potential therapeutic targets. The normalization of relative qRT-PCR data using a suitable reference gene is a crucial prerequisite for obtaining reliable conclusions. No validated housekeeping genes have been reported for the relative quantification of the mRNA expression profile activated in in-vitro ischaemic conditions, whereas for the in-vivo model different reference genes have been used.     

Proliferating mesodermal cells in murine embryos exhibiting macrophage and lymphendothelial characteristics

Background  

The data on the embryonic origin of lymphatic endothelial cells (LECs) from either deep embryonic veins or mesenchymal (or circulating) lymphangioblasts presently available remain inconsistent. In various vertebrates, markers for LECs are first expressed in specific segments of embryonic veins arguing for a venous origin of lymph vessels. Very recently, studies on the mouse have strongly supported this view. However, in the chick, we have observed a dual origin of LECs from veins and from mesodermal lymphangioblasts. Additionally, in murine embryos we have detected mesenchymal cells that co-express LEC markers and the pan-leukocyte marker CD45. Here, we have characterized the mesoderm of murine embryos with LEC markers Prox1, Lyve-1 and LA102 in combination with macrophage markers CD11b and F4/80.     

A consensus genetic map of sorghum that integrates multiple component maps and high-throughput Diversity Array Technology (DArT) markers

Background  

Sorghum genome mapping based on DNA markers began in the early 1990s and numerous genetic linkage maps of sorghum have been published in the last decade, based initially on RFLP markers with more recent maps including AFLPs and SSRs and very recently, Diversity Array Technology (DArT) markers. It is essential to integrate the rapidly growing body of genetic linkage data produced through DArT with the multiple genetic linkage maps for sorghum generated through other marker technologies. Here, we report on the colinearity of six independent sorghum component maps and on the integration of these component maps into a single reference resource that contains commonly utilized SSRs, AFLPs, and high-throughput DArT markers.     

Controlled spatial and conformational display of immobilised bone morphogenetic protein-2 and osteopontin signalling motifs regulates osteoblast adhesion and differentiation <Emphasis Type="Italic">in vitro</Emphasis>

Background  

The interfacial molecular mechanisms that regulate mammalian cell growth and differentiation have important implications for biotechnology (production of cells and cell products) and medicine (tissue engineering, prosthetic implants, cancer and developmental biology). We demonstrate here that engineered protein motifs can be robustly displayed to mammalian cells in vitro in a highly controlled manner using a soluble protein scaffold designed to self assemble on a gold surface.     

Classification of ancient mammal individuals using dental pulp MALDI-TOF MS peptide profiling

Background

The classification of ancient animal corpses at the species level remains a challenging task for forensic scientists and anthropologists. Severe damage and mixed, tiny pieces originating from several skeletons may render morphological classification virtually impossible. Standard approaches are based on sequencing mitochondrial and nuclear targets.

Methodology/Principal Findings

We present a method that can accurately classify mammalian species using dental pulp and mass spectrometry peptide profiling. Our work was organized into three successive steps. First, after extracting proteins from the dental pulp collected from 37 modern individuals representing 13 mammalian species, trypsin-digested peptides were used for matrix-assisted laser desorption/ionization time-of-flight mass spectrometry analysis. The resulting peptide profiles accurately classified every individual at the species level in agreement with parallel cytochrome b gene sequencing gold standard. Second, using a 279–modern spectrum database, we blindly classified 33 of 37 teeth collected in 37 modern individuals (89.1%). Third, we classified 10 of 18 teeth (56%) collected in 15 ancient individuals representing five mammal species including human, from five burial sites dating back 8,500 years. Further comparison with an upgraded database comprising ancient specimen profiles yielded 100% classification in ancient teeth. Peptide sequencing yield 4 and 16 different non-keratin proteins including collagen (alpha-1 type I and alpha-2 type I) in human ancient and modern dental pulp, respectively.

Conclusions/Significance

Mass spectrometry peptide profiling of the dental pulp is a new approach that can be added to the arsenal of species classification tools for forensics and anthropology as a complementary method to DNA sequencing. The dental pulp is a new source for collagen and other proteins for the species classification of modern and ancient mammal individuals.     

Proximity ligation <Emphasis Type="Italic">in situ</Emphasis> assay for monitoring the global DNA methylation in cells

Background  

DNA methylation has a central role in the epigenetic control of mammalian gene expression, and is required for X inactivation, genomics imprinting and silencing of retrotransposons and repetitive sequences. Thus, several technologies have been developed to measure the degree of DNA methylation.     

PRESTO: Rapid calculation of order statistic distributions and multiple-testing adjusted P-values via permutation for one and two-stage genetic association studies

Background  

Large-scale genetic association studies can test hundreds of thousands of genetic markers for association with a trait. Since the genetic markers may be correlated, a Bonferroni correction is typically too stringent a correction for multiple testing. Permutation testing is a standard statistical technique for determining statistical significance when performing multiple correlated tests for genetic association. However, permutation testing for large-scale genetic association studies is computationally demanding and calls for optimized algorithms and software. PRESTO is a new software package for genetic association studies that performs fast computation of multiple-testing adjusted P-values via permutation of the trait.     

Optimal Step Length EM Algorithm (OSLEM) for the estimation of haplotype frequency and its application in lipoprotein lipase genotyping

Background  

Haplotype based linkage disequilibrium (LD) mapping has become a powerful and cost-effective method for performing genetic association studies, particularly in the search for genetic markers in linkage disequilibrium with complex disease loci. Various methods (e.g. Monte-Carlo (Gibbs sampling); EM (expectation maximization); and Clark's method) have been used to estimate haplotype frequencies from routine genotyping data.     

STARNET 2: a web-based tool for accelerating discovery of gene regulatory networks using microarray co-expression data

Background  

Although expression microarrays have become a standard tool used by biologists, analysis of data produced by microarray experiments may still present challenges. Comparison of data from different platforms, organisms, and labs may involve complicated data processing, and inferring relationships between genes remains difficult.     

Three-parameter lognormal distribution ubiquitously found in cDNA microarray data and its application to parametric data treatment

Background  

To cancel experimental variations, microarray data must be normalized prior to analysis. Where an appropriate model for statistical data distribution is available, a parametric method can normalize a group of data sets that have common distributions. Although such models have been proposed for microarray data, they have not always fit the distribution of real data and thus have been inappropriate for normalization. Consequently, microarray data in most cases have been normalized with non-parametric methods that adjust data in a pair-wise manner. However, data analysis and the integration of resultant knowledge among experiments have been difficult, since such normalization concepts lack a universal standard.     

Comparison of bovine leukemia virus (BLV) and CMV promoter-driven reporter gene expression in BLV-infected and non-infected cells

Background  

Viral promoters are used in mammalian expression vectors because they generally have strong activity in a wide variety of cells of differing tissues and species.