MedWorm.com provides a medical RSS filtering service. Over 6000 RSS medical sources are combined and output via different filters. This feed contains the latest items from the 'Springer protocols feed by Bioinformatics' source. Thu, 09 Feb 2012 22:12:29 +0100 http://www.medworm.com/index.php?rid=5484132&cid=s_37118_79_f&fid=37118&url=http%3A%2F%2Fwww.springerprotocols.com%2FAbstract%2Fdoi%2F10.1007%2F978-1-61779-400-1_2 In this chapter, we demonstrate the usability of the KEGG (Kyoto encyclopedia of genes and genomes) databases and tools, especially focusing on the visualization of the omics data. The desktop application KegArray and many Web-based tools are tightly integrated with the KEGG knowledgebase, which helps visualize and interpret large amount of data derived from high-throughput measurement techniques including microarray, metagenome, and metabolome analyses. Recently developed resources for human disease, drug, and plant research are also mentioned. (Source: Springer protocols feed by Bioinformatics) Springer protocols feed by Bioinformatics news http://www.medworm.com/rss/comments.php?id=5484132 Thu, 08 Dec 2011 16:26:14 +0100 5484132 http://www.medworm.com/index.php?rid=5484131&cid=s_37118_79_f&fid=37118&url=http%3A%2F%2Fwww.springerprotocols.com%2FAbstract%2Fdoi%2F10.1007%2F978-1-61779-400-1_1 DNA microarray technology has been used for genome-wide gene expression studies that incorporate molecular genetics and computer science analyses on massive levels. The availability of microarrays permit the simultaneous analysis of tens of thousands of genes for the purposes of gene discovery, disease diagnosis, improved drug development, and therapeutics tailored to specific disease processes. In this chapter, we provide an overview on the current state of common microarray technologies and platforms. Since many genes contribute to normal functioning, research efforts are moving from the search for a disease-specific gene to the understanding of the biochemical and molecular functioning of a variety of genes whose disrupted interaction in complicated networks can lead to a disease state.... Springer protocols feed by Bioinformatics news http://www.medworm.com/rss/comments.php?id=5484131 Thu, 08 Dec 2011 16:26:14 +0100 5484131 http://www.medworm.com/index.php?rid=5418693&cid=s_37118_79_f&fid=37118&url=http%3A%2F%2Fwww.springerprotocols.com%2FAbstract%2Fdoi%2F10.1007%2F978-1-61779-412-4_23 The rapidly emerging ability to design and construct synthetic gene networks in mammalian cells is based on the availability of mutually compatible genetic switches that enable the time-dependent induction of transgene expression in response to the dose of an externally applied stimulus. As these genetic switches are inherently compatible with mammalian cell physiology, they are as well predestined to control the functionality of cell-free synthetic devices within an overall physiologic background. In this chapter, we describe how a genetic switch that was originally designed for gene therapeutic studies can be applied in materials science to design and construct a biohybrid hydrogel that can be used to release a therapeutic growth factor in response to an externally applied stimulus for c... Springer protocols feed by Bioinformatics news http://www.medworm.com/rss/comments.php?id=5418693 Fri, 18 Nov 2011 06:01:32 +0100 5418693 http://www.medworm.com/index.php?rid=5418692&cid=s_37118_79_f&fid=37118&url=http%3A%2F%2Fwww.springerprotocols.com%2FAbstract%2Fdoi%2F10.1007%2F978-1-61779-412-4_22 Advances in the development of molecular tools for the inducible control of transcription, translation, and protein degradation are the basis for the rapidly emerging design and construction of synthetic gene networks in mammalian cells. (Source: Springer protocols feed by Bioinformatics) Springer protocols feed by Bioinformatics news http://www.medworm.com/rss/comments.php?id=5418692 Fri, 18 Nov 2011 06:01:32 +0100 5418692 http://www.medworm.com/index.php?rid=5418691&cid=s_37118_79_f&fid=37118&url=http%3A%2F%2Fwww.springerprotocols.com%2FAbstract%2Fdoi%2F10.1007%2F978-1-61779-412-4_21 Synthetic biology methods are routinely applied in the plant field as in other eukaryotic model systems. Several synthetic components have been developed in plants and an increasing number of studies report on the assembly into functional synthetic genetic circuits. This chapter gives an overview of the existing plant genetic networks and describes in detail the application of two systems for inducible gene expression. The ethanol-inducible system relies on the ethanol-responsive interaction of the AlcA transcriptional activator and the AlcR receptor resulting in the transcription of the gene of interest (GOI). In comparison, the translational fusion of GOI and the glucocorticoid receptor (GR) domain leads to the dexamethasone-dependent nuclear translocation of the GOI::GR protein. This ch... Springer protocols feed by Bioinformatics news http://www.medworm.com/rss/comments.php?id=5418691 Fri, 18 Nov 2011 06:01:32 +0100 5418691 http://www.medworm.com/index.php?rid=5418690&cid=s_37118_79_f&fid=37118&url=http%3A%2F%2Fwww.springerprotocols.com%2FAbstract%2Fdoi%2F10.1007%2F978-1-61779-412-4_20 Synthetic biology is an engineering approach to biology. A synthetic biologist wants to describe biological molecules and their subdomains as well-defined parts of a molecular machine. To achieve this goal, synthetic biologists rebuild minimal functional biological systems from well-defined parts or they design new molecules that do not exist in nature but have new and useful functions. In short, these engineering approaches can be summarized as “rebuild, alter, and understand.” The Drosophila S2 Schneider cell is a useful tool for both rebuilding and redesigning approaches. S2 cells are phagocytic cells that easily take up large amounts of DNA from the cell culture. They, thus, have a high cotransfection rate, allowing the coexpression of up to 12 different proteins. We have d... Springer protocols feed by Bioinformatics news http://www.medworm.com/rss/comments.php?id=5418690 Fri, 18 Nov 2011 06:01:32 +0100 5418690 http://www.medworm.com/index.php?rid=5418689&cid=s_37118_79_f&fid=37118&url=http%3A%2F%2Fwww.springerprotocols.com%2FAbstract%2Fdoi%2F10.1007%2F978-1-61779-412-4_19 A major focus in synthetic biology is the rational design and implementation of gene circuits to control dynamics of individual cells and, increasingly, cellular populations. Population-level control is highlighted in recent studies which attempt to design and implement synthetic ecosystems (or engineered microbial consortia). On the one hand, these engineered systems may serve as a critical technological foundation for practical applications. On the other hand, they may serve as well-defined model systems to examine biological questions of broad relevance. Here, using a synthetic predator–prey ecosystem as an example, we illustrate the basic experimental techniques involved in system implementation and characterization. By extension, these techniques are applicable to the analysis o... Springer protocols feed by Bioinformatics news http://www.medworm.com/rss/comments.php?id=5418689 Fri, 18 Nov 2011 06:01:32 +0100 5418689 http://www.medworm.com/index.php?rid=5418688&cid=s_37118_79_f&fid=37118&url=http%3A%2F%2Fwww.springerprotocols.com%2FAbstract%2Fdoi%2F10.1007%2F978-1-61779-412-4_18 Most microorganisms in nature subsist as heterogeneous surface-associated communities called biofilms. In biofilms members of one or more microbial species live together for multiple generations, and this allows them to cooperate and co-adapt. The ability to reliably manipulate, characterize, and engineer microbial biofilms will enable controlled studies of ecosystem dynamics and unprecedented design opportunities for biological sensors and actuators. Biofilms can be grown in the laboratory, and spatial structure, gene expression, and productivity (total biomass accumulation) can be observed and quantified as a function of time using confocal laser scanning microscopy. This chapter details the materials and methods necessary to grow and study engineered microbial communities in biofilms. (... Springer protocols feed by Bioinformatics news http://www.medworm.com/rss/comments.php?id=5418688 Fri, 18 Nov 2011 06:01:32 +0100 5418688 http://www.medworm.com/index.php?rid=5418687&cid=s_37118_79_f&fid=37118&url=http%3A%2F%2Fwww.springerprotocols.com%2FAbstract%2Fdoi%2F10.1007%2F978-1-61779-412-4_17 Bacterial synthetic gene networks are constructed by manipulating the regulation of genes inside a cell, with the purpose of eliciting novel regulatory behaviors. The methods for manipulating genes and gene regulation in E. coli are well established, making it the preferred host for basic studies of synthetic networks. We focus our work on constructing two kinds of synthetic gene networks: toggle switches (bistable systems) and oscillators. Toggle switches are capable of exhibiting two stable steady states of gene expression (OFF and ON) without stable intermediate states; the steady state reached by the system depends on the previous history of the system. Biological oscillators exhibit regular cycles in gene expression around an unstable steady state. Studying these two kinds of syntheti... Springer protocols feed by Bioinformatics news http://www.medworm.com/rss/comments.php?id=5418687 Fri, 18 Nov 2011 06:01:32 +0100 5418687 http://www.medworm.com/index.php?rid=5418686&cid=s_37118_79_f&fid=37118&url=http%3A%2F%2Fwww.springerprotocols.com%2FAbstract%2Fdoi%2F10.1007%2F978-1-61779-412-4_16 A good part of the contemporary synthetic biology agenda aims at reprogramming microorganisms to enhance existing functions and/or perform new tasks. Moreover, the functioning of complex regulatory networks, or even a single gene, is revealed only when perturbations are entered in the corresponding dynamic systems and the outcome monitored. These endeavors rely on the availability of genetic tools to successfully modify á la carte the chromosome of target bacteria. Key aspects to this end include the removal of undesired genomic segments, systems for the production of directed mutants and allelic replacements, random mutant libraries to discover new functions, and means to stably implant larger genetic networks into the genome of specific hosts. The list of gram-negative species tha... Springer protocols feed by Bioinformatics news http://www.medworm.com/rss/comments.php?id=5418686 Fri, 18 Nov 2011 06:01:32 +0100 5418686 http://www.medworm.com/index.php?rid=5418685&cid=s_37118_79_f&fid=37118&url=http%3A%2F%2Fwww.springerprotocols.com%2FAbstract%2Fdoi%2F10.1007%2F978-1-61779-412-4_15 Here, we document a technique to reduce the size of the genome of Pseudomonas putida by using a combinatorial mini-Tn5-targeted Flp-FRT recombination system. This method combines random insertions with the site-specific Flp-FRT recombination system to generate successive random deletions in a single strain in which parts of the genome are excised via the action of the cognate flippase. For this purpose, we have generated two mini-Tn5 transposon mutant libraries with single and double integrations of either mini-Tn5 KpF alone or mini-Tn5 KpF in parallel with mini-Tn5 TF, respectively. These mini-Tn5 transposons carry different selectable markers and each has an FRT (Flippase Recognition Target) site. Mapping of the position of both mini-Tn5 transposons in the chromosome of P. putida was con... Springer protocols feed by Bioinformatics news http://www.medworm.com/rss/comments.php?id=5418685 Fri, 18 Nov 2011 06:01:32 +0100 5418685 http://www.medworm.com/index.php?rid=5418684&cid=s_37118_79_f&fid=37118&url=http%3A%2F%2Fwww.springerprotocols.com%2FAbstract%2Fdoi%2F10.1007%2F978-1-61779-412-4_14 Cellular hosts are widely used for the production of chemical compounds, including pharmaceutics, fuels, and specialty chemicals. However, common metabolic engineering techniques are limited in their capacity to elicit multigenic, complex phenotypes. These phenotypes can include non-pathway-based traits, such as tolerance and productivity. Global transcription machinery engineering (gTME) is a generic methodology for eliciting these complex cellular phenotypes. In gTME, dominant mutant alleles of a transcription-related protein are screened for their ability to reprogram cellular metabolism and regulation, resulting in a unique and desired phenotype. gTME has been successfully applied to both prokaryotic and eukaryotic systems, resulting in improved environmental tolerances, metabolite pro... Springer protocols feed by Bioinformatics news http://www.medworm.com/rss/comments.php?id=5418684 Fri, 18 Nov 2011 06:01:32 +0100 5418684 http://www.medworm.com/index.php?rid=5418683&cid=s_37118_79_f&fid=37118&url=http%3A%2F%2Fwww.springerprotocols.com%2FAbstract%2Fdoi%2F10.1007%2F978-1-61779-412-4_9 Within the last few years, a set of synthetic riboswitches has been engineered, which expands the toolbox of genetic regulatory devices. Small molecule binding aptamers have been used for the design of such riboswitches by insertion into untranslated regions of mRNAs, exploiting the fact that upon ligand binding the RNA structure interferes either with translation initiation or pre-mRNA splicing in yeast. In combination with self-cleaving ribozymes, aptamers have been used to modulate RNA stability. In this chapter, we discuss the applicability of different aptamers, ways to identify novel genetic devices, the pros and cons of various insertion sites and the application of allosteric ribozymes. Our expertise help to apply synthetic riboswitches to engineer complex genetic circuits. (Source... Springer protocols feed by Bioinformatics news http://www.medworm.com/rss/comments.php?id=5418683 Fri, 18 Nov 2011 06:01:32 +0100 5418683 http://www.medworm.com/index.php?rid=5418682&cid=s_37118_79_f&fid=37118&url=http%3A%2F%2Fwww.springerprotocols.com%2FAbstract%2Fdoi%2F10.1007%2F978-1-61779-412-4_8 Customized zinc-finger nucleases (ZFNs) have developed into a promising technology to precisely alter mammalian genomes for biomedical research, biotechnology, or human gene therapy. In the context of synthetic biology, the targeted integration of a transgene or reporter cassette into a “neutral site” of the human genome, such as the AAVS1 locus, permits the generation of isogenic human cell lines with two major advantages over standard genetic manipulation techniques: minimal integration site-dependent effects on the transgene and, vice versa, no functional perturbation of the host-cell transcriptome. Here we describe in detail how ZFNs can be employed to target integration of a transgene cassette into the AAVS1 locus and how to characterize the targeted cells by PCR-based gen... Springer protocols feed by Bioinformatics news http://www.medworm.com/rss/comments.php?id=5418682 Fri, 18 Nov 2011 06:01:32 +0100 5418682 http://www.medworm.com/index.php?rid=5418681&cid=s_37118_79_f&fid=37118&url=http%3A%2F%2Fwww.springerprotocols.com%2FAbstract%2Fdoi%2F10.1007%2F978-1-61779-412-4_7 Protein–protein interactions are crucial for the vast majority of biological processes. To fully understand these processes therefore requires methods for identifying protein interactions within the complex cellular environment. To isolate interacting proteins, we have developed a simple and reliable genetic selection by exploiting the inbuilt “hitchhiker” mechanism of the Escherichia coli twin-arginine translocation (Tat) pathway. This method is based on the unique ability of the Tat system to efficiently co-localize noncovalent complexes of two folded polypeptides to the periplasmic space of E. coli. The genetic selection is comprised of two engineered fusion proteins: an N-terminal Tat signal peptide fused to the protein of interest, and the known or putative partner p... Springer protocols feed by Bioinformatics news http://www.medworm.com/rss/comments.php?id=5418681 Fri, 18 Nov 2011 06:01:32 +0100 5418681 http://www.medworm.com/index.php?rid=5418680&cid=s_37118_79_f&fid=37118&url=http%3A%2F%2Fwww.springerprotocols.com%2FAbstract%2Fdoi%2F10.1007%2F978-1-61779-412-4_1 Computational synthetic biology has borrowed methods, concepts, and techniques from systems biology and electrical engineering. Features of tools for the analysis of biochemical networks and the design of electric circuits have been combined to develop new software, where Standard Biological Parts (physically stored at the MIT Registry) have a mathematical description, based on mass action or Hill kinetics, and can be assembled into genetic networks in a visual, “drag & drop” fashion. Recent tools provide the user with databases, simulation environments, formal languages, and even algorithms for circuit automatic design to refine and speed up gene network construction. Moreover, advances in automation of DNA assembly indicate that synthetic biology software soon will drive ... Springer protocols feed by Bioinformatics news http://www.medworm.com/rss/comments.php?id=5418680 Fri, 18 Nov 2011 06:01:32 +0100 5418680 http://www.medworm.com/index.php?rid=5418679&cid=s_37118_79_f&fid=37118&url=http%3A%2F%2Fwww.springerprotocols.com%2FAbstract%2Fdoi%2F10.1007%2F978-1-61779-412-4_3 In engineering, the use of mathematical modeling for design purposes has a long history. Long before any technical realization, a system is planned, simulated, and tested extensively on the computer. In biosciences, however, the application of model-based design before going to the wet lab is still rather rare but has particularly high potential in synthetic biology. We demonstrate exemplarily how mathematical modeling and numerical optimization can be used for the design of a circadian rhythm that is supposed to oscillate robustly with respect to uncertainty in system parameters. (Source: Springer protocols feed by Bioinformatics) Springer protocols feed by Bioinformatics news http://www.medworm.com/rss/comments.php?id=5418679 Fri, 18 Nov 2011 06:01:32 +0100 5418679 http://www.medworm.com/index.php?rid=5418678&cid=s_37118_79_f&fid=37118&url=http%3A%2F%2Fwww.springerprotocols.com%2FAbstract%2Fdoi%2F10.1007%2F978-1-61779-412-4_2 Synthetic Biology is founded on the idea that complex biological systems are built most effectively when the task is divided in abstracted layers and all required components are readily available and well-described. This requires interdisciplinary collaboration at several levels and a common understanding of the functioning of each component. Standardization of the physical composition and the description of each part is required as well as a controlled vocabulary to aid design and ensure interoperability. Here, we describe standardization initiatives from several disciplines, which can contribute to Synthetic Biology. We provide examples of the concerted standardization efforts of the BioBricks Foundation comprising the request for comments (RFC) and the Registry of Standardized Biologica... Springer protocols feed by Bioinformatics news http://www.medworm.com/rss/comments.php?id=5418678 Fri, 18 Nov 2011 06:01:32 +0100 5418678 http://www.medworm.com/index.php?rid=5418677&cid=s_37118_79_f&fid=37118&url=http%3A%2F%2Fwww.springerprotocols.com%2FAbstract%2Fdoi%2F10.1007%2F978-1-61779-412-4_4 Synthetic biology aims at designing and building new biological functions in living organisms. The complexity of cellular regulation (regulatory, metabolic, and signaling interactions, and their coordinated action) can be tackled via the development of quantitative mathematical models. These models are useful to test biological hypotheses and observations, and to predict the possible behaviors of a synthetic network. Indeed, synthetic biology uses such models to design synthetic networks, prior to their construction in the cell, to perform specific tasks, or to change a biological process in a desired way. The synthetic network is built by assembling biological “parts” taken from different systems; therefore it is fundamental to identify, isolate, and test regulatory motifs whi... Springer protocols feed by Bioinformatics news http://www.medworm.com/rss/comments.php?id=5418677 Fri, 18 Nov 2011 06:01:32 +0100 5418677 http://www.medworm.com/index.php?rid=5418676&cid=s_37118_79_f&fid=37118&url=http%3A%2F%2Fwww.springerprotocols.com%2FAbstract%2Fdoi%2F10.1007%2F978-1-61779-412-4_10 One of the longstanding challenges in synthetic biology is rational design of complex regulatory circuitry with multiple biological inputs, complex internal processing, and physiologically active outputs. We have previously proposed how to address this challenge in the case of transcription factor inputs. Here we describe the methods used to construct these synthetic circuits, capable of performing logic integration of transcription factor inputs using microRNA expression vectors and RNA interference (RNAi). The circuits operate in mammalian cells and they can serve as starting point for more complex synthetic information processing networks in these cells. (Source: Springer protocols feed by Bioinformatics) Springer protocols feed by Bioinformatics news http://www.medworm.com/rss/comments.php?id=5418676 Fri, 18 Nov 2011 06:01:32 +0100 5418676 http://www.medworm.com/index.php?rid=5418675&cid=s_37118_79_f&fid=37118&url=http%3A%2F%2Fwww.springerprotocols.com%2FAbstract%2Fdoi%2F10.1007%2F978-1-61779-412-4_11 An important basic requirement of synthetic genetic networks is the option of external control of gene expression. Although several chemically inducible systems are available, all of these suffer from the common problem: the chemical inducers are difficult to remove so that to terminate the response. We have described a regulatory expression system for yeast, which employs light as inducer. This light switch translates light-controlled protein–protein interactions into the transcription of selected genes in a dose-dependent and reversible manner. (Source: Springer protocols feed by Bioinformatics) Springer protocols feed by Bioinformatics news http://www.medworm.com/rss/comments.php?id=5418675 Fri, 18 Nov 2011 06:01:32 +0100 5418675 http://www.medworm.com/index.php?rid=5418674&cid=s_37118_79_f&fid=37118&url=http%3A%2F%2Fwww.springerprotocols.com%2FAbstract%2Fdoi%2F10.1007%2F978-1-61779-412-4_12 Remote control of cells is a desirable feature in synthetic biology. We established a light-switchable interfering peptide (iPEP) which controls gene expression by modulating the activity of a transcription factor. For photo-switching, the iPEP is cross-linked with a cis-trans isomerizable cross-linker in such a way that the light-activated cis form enables inhibitor folding rendering it active, whereas the dark-adapted trans form forces the inhibitor into an inactive form. Switching can be repeated in both directions. The iPEP acts as dominant-negative inhibitor targeting c-Jun and c-Fos of the transcription factor activator protein-1 (AP-1). Light-activated peptides exhibited much stronger inhibition of AP-1:DNA complexes and interference with gene transcription than their nonactivated c... Springer protocols feed by Bioinformatics news http://www.medworm.com/rss/comments.php?id=5418674 Fri, 18 Nov 2011 06:01:32 +0100 5418674 http://www.medworm.com/index.php?rid=5418673&cid=s_37118_79_f&fid=37118&url=http%3A%2F%2Fwww.springerprotocols.com%2FAbstract%2Fdoi%2F10.1007%2F978-1-61779-412-4_13 Techniques to manipulate cellular gene expression such that amino acid analogs not encoded by the genetic code are incorporated into a polypeptide chain have recently gained increasing interest. The so-called noncanonical amino acids often have unusual properties that can be translated into target proteins by reprogrammed ribosomal protein synthesis. Residue-specific substitution of a specific canonical amino acid by its analogs provokes global effects in the resulting protein congeners that include improved stability or catalytic activity, reduced redox sensitivity, as well as altered spectral properties. Thus, the approach holds great promise for the engineering of synthetic proteins. (Source: Springer protocols feed by Bioinformatics) Springer protocols feed by Bioinformatics news http://www.medworm.com/rss/comments.php?id=5418673 Fri, 18 Nov 2011 06:01:32 +0100 5418673 http://www.medworm.com/index.php?rid=5418672&cid=s_37118_79_f&fid=37118&url=http%3A%2F%2Fwww.springerprotocols.com%2FAbstract%2Fdoi%2F10.1007%2F978-1-61779-412-4_5 Late human immunodeficiency virus (HIV)-derived RNAs encoding relevant therapeutic targets or promising vaccine compounds, such as the HIV-1 group-specific antigen (Gag), are translocated from the nucleus into the cytoplasm via sophisticated export machinery. Relevant steps include the concerted action of several cis-acting RNA elements with the viral Rev-shuttle protein and several cellular components (Ran1/Exportin; Crm1). Based on detailed understanding of the molecular mechanisms guiding this complex process, we used rational codon usage modification to design and reprogram a GFP encoding reporter RNA now exactly mimicking the complex transcriptional and posttranscriptional regulation of late lentiviral mRNAs. (Source: Springer protocols feed by Bioinformatics) Springer protocols feed by Bioinformatics news http://www.medworm.com/rss/comments.php?id=5418672 Fri, 18 Nov 2011 06:01:32 +0100 5418672 http://www.medworm.com/index.php?rid=5418671&cid=s_37118_79_f&fid=37118&url=http%3A%2F%2Fwww.springerprotocols.com%2FAbstract%2Fdoi%2F10.1007%2F978-1-61779-412-4_6 Characterizing libraries of mutant proteins is a challenging task, but can lead to detailed functional insights on a specific protein, and general insights for families of proteins such as transcription factors. Challenges in mutant protein screening consist in synthesizing the necessary expression-ready DNA constructs and transforming them into a suitable host for protein expression. Protein purification and characterization are also non-trivial tasks that are not easily scalable to hundreds or thousands of protein variants. Here we describe a method based on a high-throughput microfluidic platform to screen and characterize the binding profile of hundreds of transcription factor variants. DNA constructs are synthesized by a rapid two-step PCR approach without the need of cloning or trans... Springer protocols feed by Bioinformatics news http://www.medworm.com/rss/comments.php?id=5418671 Fri, 18 Nov 2011 06:01:32 +0100 5418671 http://www.medworm.com/index.php?rid=5179401&cid=s_37118_79_f&fid=37118&url=http%3A%2F%2Fwww.springerprotocols.com%2FAbstract%2Fdoi%2F10.1007%2F978-1-61779-276-2_9 Genome sequencing efforts have reformed the nature of biological inquiry, prompting the development of technologies for the functional annotation of mammalian genes. Based on methodologies originally discovered in plants and Caenorhabditis elegans, RNA interference has offered cell biologists an effective and reproducible approach to perturb gene function in mammalian cells and whole organisms. Initial application of RNA interference libraries targeting the human and mouse genomes relied on arrayed screening approaches, whereby each unique RNA interference reagent is arrayed into individual wells of a microtiter plate. These screens are not trivial to perform, requiring a substantial investment in infrastructure. In the past decade, many technological advances have been made that make geno... Springer protocols feed by Bioinformatics news http://www.medworm.com/rss/comments.php?id=5179401 Wed, 31 Aug 2011 05:11:36 +0100 5179401 http://www.medworm.com/index.php?rid=5179400&cid=s_37118_79_f&fid=37118&url=http%3A%2F%2Fwww.springerprotocols.com%2FAbstract%2Fdoi%2F10.1007%2F978-1-61779-276-2_8 High-throughput methodologies have created new opportunities for studying biological phenomena in an unbiased manner. Using automated cell manipulations and microscopy platforms, it is now possible to easily screen entire genomes for genes that affect any cellular process that can be visualized. The onset of these methodologies promises that the near future will bring with it a more comprehensive and richly integrated understanding of complex and dynamic cellular structures and processes. In this review, we describe how to couple systematic genetic tools in the budding yeast Saccharomyces cerevisiae alongside robotic visualization systems to attack biological questions. The combination of high-throughput microscopy screens with the powerful, yet simple, yeast model system for studying the ... Springer protocols feed by Bioinformatics news http://www.medworm.com/rss/comments.php?id=5179400 Wed, 31 Aug 2011 05:11:35 +0100 5179400 http://www.medworm.com/index.php?rid=5179399&cid=s_37118_79_f&fid=37118&url=http%3A%2F%2Fwww.springerprotocols.com%2FAbstract%2Fdoi%2F10.1007%2F978-1-61779-276-2_7 Cellular processes are carried out through a series of molecular interactions. Various experimental approaches can be used to investigate these functional relationships on a large-scale. Recently, the power of investigating biological systems from the perspective of genetic (gene–gene, or epistatic) interactions has been evidenced by the ability to elucidate novel functional relationships. Examples of functionally related genes include genes that buffer each other’s function or impinge on the same biological process. Genetic interactions have traditionally been investigated in bacteria by combining pairs of mutations (for example, gene deletions) and assessing deviation of the phenotype of each double mutant from an expected neutral (or no interaction) phenotype. Fitness is a p... Springer protocols feed by Bioinformatics news http://www.medworm.com/rss/comments.php?id=5179399 Wed, 31 Aug 2011 05:11:34 +0100 5179399 http://www.medworm.com/index.php?rid=5179398&cid=s_37118_79_f&fid=37118&url=http%3A%2F%2Fwww.springerprotocols.com%2FAbstract%2Fdoi%2F10.1007%2F978-1-61779-276-2_6 Peptide recognition modules (PRMs) play critical roles in cellular processes, including differentiation, proliferation and cytoskeleton organization. PRMs normally bind to short linear motifs in protein ligands, and by so doing recruit proteins into signaling complexes. Based on the binding specificity profile of a PRM, one can predict putative natural interaction partners by searching genome databases. Candidate interaction partners can in turn provide clues to assemble potential in vivo protein complexes that the PRM may be involved with. Combinatorial peptide libraries have proven to be effective tools for profiling the binding specificities of PRMs. Herein, we describe high-throughput methods for the expression and purification of PRM proteins and the use of peptide-phage libraries for... Springer protocols feed by Bioinformatics news http://www.medworm.com/rss/comments.php?id=5179398 Wed, 31 Aug 2011 05:11:34 +0100 5179398 http://www.medworm.com/index.php?rid=5179397&cid=s_37118_79_f&fid=37118&url=http%3A%2F%2Fwww.springerprotocols.com%2FAbstract%2Fdoi%2F10.1007%2F978-1-61779-276-2_5 Multiprotein complexes are essential building blocks for many cellular processes in an organism. Taking the process of transcription as an example, the interplay of several chromatin-remodeling complexes is responsible for the tight regulation of gene expression. Knowing how those proteins associate into protein complexes not only helps to improve our understanding of these cellular processes, but can also lead to the discovery of the function of novel interacting proteins. Given the large number of proteins with little to no functional annotation throughout many organisms, including human, the identification and characterization of protein complexes has grown into a major focus of network biology. Toward this goal, we have developed several computational approaches based upon label-free q... Springer protocols feed by Bioinformatics news http://www.medworm.com/rss/comments.php?id=5179397 Wed, 31 Aug 2011 05:11:34 +0100 5179397 http://www.medworm.com/index.php?rid=5179396&cid=s_37118_79_f&fid=37118&url=http%3A%2F%2Fwww.springerprotocols.com%2FAbstract%2Fdoi%2F10.1007%2F978-1-61779-276-2_4 Proteome-wide analysis of protein C-termini has long been inaccessible, but is now enabled by a newly developed negative selection strategy we term C-terminomics. In this procedure, amine- and carboxyl groups of full-length proteins are chemically protected. After trypsin digestion, N-terminal and internal tryptic peptides – but not C-terminal peptides – posses newly formed, unprotected C-termini that are removed by coupling to the high-molecular-weight polymer poly-allylamine. Ultrafiltration separates the uncoupled, blocked C-terminal peptides that are subsequently analyzed by liquid chromatography-tandem mass spectrometry. On a proteome-wide scale, this strategy profiles native protein C-termini together with neo C-termini generated by endoproteolytic cleavage or processive ... Springer protocols feed by Bioinformatics news http://www.medworm.com/rss/comments.php?id=5179396 Wed, 31 Aug 2011 05:11:33 +0100 5179396 http://www.medworm.com/index.php?rid=5179395&cid=s_37118_79_f&fid=37118&url=http%3A%2F%2Fwww.springerprotocols.com%2FAbstract%2Fdoi%2F10.1007%2F978-1-61779-276-2_3 Secretagogin is a calcium-binding protein whose expression is characterised in neuroendocrine, pancreatic, and retinal cells. We have used an array-based proteomic approach with the prokaryotically expressed human protein array (hEx1) and the eukaryotically expressed human protein array (Protoarray) to identify novel calcium-regulated interaction networks of secretagogin. Screening of these arrays with fluorophore-labelled secretagogin in the presence of Ca2+ ions led to the identification of 12 (hEx1) and 6 (Protoarray) putative targets. A number of targets were identified in both array screens. The putative targets from the hEx1 array were expressed, purified, and subjected to binding analysis using surface plasmon resonance. This identified binding affinities for nine novel secretagogin... Springer protocols feed by Bioinformatics news http://www.medworm.com/rss/comments.php?id=5179395 Wed, 31 Aug 2011 05:11:32 +0100 5179395 http://www.medworm.com/index.php?rid=5179394&cid=s_37118_79_f&fid=37118&url=http%3A%2F%2Fwww.springerprotocols.com%2FAbstract%2Fdoi%2F10.1007%2F978-1-61779-276-2_2 Protein complexes and protein–protein interactions (PPIs) are fundamental for most biological functions. Deciphering the extensive protein interaction networks that occur within cellular contexts has become a logical extension to the human genome project. Proteome-scale interactome analysis of mammalian systems requires efficient methods for accurately detecting PPIs with specific considerations for the intrinsic technical challenges of mammalian genome manipulation. In this chapter, we outline in detail an innovative lentiviral-based functional proteomic approach that can be used to rapidly characterize protein complexes from a broad range of mammalian cell lines. This method integrates the following key features: (1) lentiviral elements for efficient delivery of tagged constructs i... Springer protocols feed by Bioinformatics news http://www.medworm.com/rss/comments.php?id=5179394 Wed, 31 Aug 2011 05:11:32 +0100 5179394 http://www.medworm.com/index.php?rid=5179393&cid=s_37118_79_f&fid=37118&url=http%3A%2F%2Fwww.springerprotocols.com%2FAbstract%2Fdoi%2F10.1007%2F978-1-61779-276-2_21 Mathematical and computational models have become indispensable tools for integrating and interpreting heterogeneous biological data, understanding fundamental principles of biological system functions, genera­ting reliable testable hypotheses, and identifying potential diagnostic markers and therapeutic targets. Thus, such tools are now routinely used in the theoretical and experimental systematic investigation of biological system dynamics. Here, we discuss model building as an essential part of the theoretical and experimental analysis of biomolecular network dynamics. Specifically, we describe a procedure for defining kinetic equations and parameters of biomolecular processes, and we illustrate the use of fractional activity functions for modeling gene expression regulation by sing... Springer protocols feed by Bioinformatics news http://www.medworm.com/rss/comments.php?id=5179393 Wed, 31 Aug 2011 05:11:31 +0100 5179393 http://www.medworm.com/index.php?rid=5179392&cid=s_37118_79_f&fid=37118&url=http%3A%2F%2Fwww.springerprotocols.com%2FAbstract%2Fdoi%2F10.1007%2F978-1-61779-276-2_20 A large number of genome-scale networks, including protein–protein and genetic interaction networks, are now available for several organisms. In parallel, many studies have focused on analyzing, characterizing, and modeling these networks. Beyond investigating the topological characteristics such as degree distribution, clustering coefficient, and average shortest-path distance, another area of particular interest is the prediction of nodes (genes) with a given characteristic (labels) – for example prediction of genes that cause a particular phenotype or have a given function. In this chapter, we describe methods and algorithms for predicting node labels from network-based datasets with an emphasis on label propagation algorithms (LPAs) and their relation to local neighborhood ... Springer protocols feed by Bioinformatics news http://www.medworm.com/rss/comments.php?id=5179392 Wed, 31 Aug 2011 05:11:31 +0100 5179392 http://www.medworm.com/index.php?rid=5179391&cid=s_37118_79_f&fid=37118&url=http%3A%2F%2Fwww.springerprotocols.com%2FAbstract%2Fdoi%2F10.1007%2F978-1-61779-276-2_1 The yeast two-hybrid (Y2H) system is a powerful tool to identify binary protein–protein interactions. Here, we describe array-based two-hybrid methods that use defined libraries of open reading frames (ORFs) and pooled prey library screenings that use random genomic or cDNA libraries. The array-based Y2H system is well-suited for interactome studies of existing ORFeomes or subsets thereof, preferentially in a recombination-based cloning system. Array-based Y2H screens efficiently reduce false positives by using built-in controls, retesting, and evaluation of background activation. Hands-on time and the amount of used resources grow exponentially with the number of tested proteins; this is a disadvantage for large genome sizes. For large genomes, random library screen may be more effi... Springer protocols feed by Bioinformatics news http://www.medworm.com/rss/comments.php?id=5179391 Wed, 31 Aug 2011 05:11:30 +0100 5179391 http://www.medworm.com/index.php?rid=5179390&cid=s_37118_79_f&fid=37118&url=http%3A%2F%2Fwww.springerprotocols.com%2FAbstract%2Fdoi%2F10.1007%2F978-1-61779-276-2_19 To understand the workings of the living cell, we need to characterize protein assemblies that constitute the cell (for example, the ribosome, 26S proteasome, and the nuclear pore complex). A reliable high-resolution structural characterization of these assemblies is frequently beyond the reach of current experimental methods, such as X-ray crystallography, NMR spectroscopy, electron microscopy, footprinting, chemical cross-linking, FRET spectroscopy, small-angle X-ray scattering, and proteomics. However, the information garnered from different methods can be combined and used to build computational models of the assembly structures that are consistent with all of the available datasets. Here, we describe a protocol for this integration, whereby the information is converted to a set of spa... Springer protocols feed by Bioinformatics news http://www.medworm.com/rss/comments.php?id=5179390 Wed, 31 Aug 2011 05:11:30 +0100 5179390 http://www.medworm.com/index.php?rid=5179389&cid=s_37118_79_f&fid=37118&url=http%3A%2F%2Fwww.springerprotocols.com%2FAbstract%2Fdoi%2F10.1007%2F978-1-61779-276-2_18 Cytoscape is an open-source software package that is widely used to integrate and visualize diverse data sets in biology. This chapter explains how to use Cytoscape to integrate open-source chemical information with a biological network. By visualizing information about known compound–target interactions in the context of a biological network of interest, one can rapidly identify novel avenues to perturb the system with compounds and, for example, potentially identify therapeutically relevant targets. Herein, two different protocols are explained in detail, with no prior knowledge of Cytoscape assumed, which demonstrate how to incorporate data from the ChEMBL database with either a gene–gene or a protein–protein interaction network. ChEMBL is a very large, open-source rep... Springer protocols feed by Bioinformatics news http://www.medworm.com/rss/comments.php?id=5179389 Wed, 31 Aug 2011 05:11:30 +0100 5179389 http://www.medworm.com/index.php?rid=5179388&cid=s_37118_79_f&fid=37118&url=http%3A%2F%2Fwww.springerprotocols.com%2FAbstract%2Fdoi%2F10.1007%2F978-1-61779-276-2_17 Mapping epistatic (or genetic) interactions has emerged as an important network biology approach for establishing functional relationships among genes and proteins. Epistasis networks are complementary to physical protein interaction networks, providing valuable insight into both the function of individual genes and the overall wiring of the cell. A high-throughput method termed “epistatic mini array profiles” (E-MAPs) was recently developed in yeast to quantify alleviating or aggravating interactions between gene pairs. The typical output of an E-MAP experiment is a large symmetric matrix of interaction scores. One problem with this data is the large amount of missing values – interactions that cannot be measured during the high-throughput process or whose measurements w... Springer protocols feed by Bioinformatics news http://www.medworm.com/rss/comments.php?id=5179388 Wed, 31 Aug 2011 05:11:29 +0100 5179388 http://www.medworm.com/index.php?rid=5179387&cid=s_37118_79_f&fid=37118&url=http%3A%2F%2Fwww.springerprotocols.com%2FAbstract%2Fdoi%2F10.1007%2F978-1-61779-276-2_16 Here, we present a detailed method for generating a dynamic transcriptional regulatory network from large-scale chromatin immunoprecipitation data, and functional analysis of participating factors through the identification and characterization of significantly overrepresented multi-input motifs in the network. This is done by visualizing interactive data using a network analysis tool, such as Cytoscape, clustering DNA targets of the transcription factors based on their network topologies, and statistically analyzing each cluster based on its size and properties of its members. These analyses yield testable predictions about the conditional and cooperative functions of the factors. This is a versatile approach that allows the visualization of network architecture on a genome-wide level and... Springer protocols feed by Bioinformatics news http://www.medworm.com/rss/comments.php?id=5179387 Wed, 31 Aug 2011 05:11:29 +0100 5179387 http://www.medworm.com/index.php?rid=5179386&cid=s_37118_79_f&fid=37118&url=http%3A%2F%2Fwww.springerprotocols.com%2FAbstract%2Fdoi%2F10.1007%2F978-1-61779-276-2_15 Molecular expression patterns have often been used for patient classification in oncology in an effort to improve prognostic prediction and treatment compatibility. This effort is, however, hampered by the highly heterogeneous data often seen in the molecular analysis of cancer. The lack of overall similarity between expression profiles makes it difficult to partition data using conventional data mining tools. In this chapter, the authors introduce a bioinformatics protocol that uses REACTOME pathways and patient–protein network structure (also called topology) as the basis for patient classification. (Source: Springer protocols feed by Bioinformatics) Springer protocols feed by Bioinformatics news http://www.medworm.com/rss/comments.php?id=5179386 Wed, 31 Aug 2011 05:11:29 +0100 5179386 http://www.medworm.com/index.php?rid=5179385&cid=s_37118_79_f&fid=37118&url=http%3A%2F%2Fwww.springerprotocols.com%2FAbstract%2Fdoi%2F10.1007%2F978-1-61779-276-2_14 Rarely acting in isolation, it is invariably the physical associations among proteins that define their biological activity, necessitating the study of the cellular meshwork of protein–protein interactions (PPI) before a full appreciation of gene function can be achieved. The past few years have seen a marked expansion in the both the sheer volume and number of organisms for which high-quality interaction data is available, with high-throughput interaction screening and detection techniques showing consistent improvement both in scale and sensitivity. Although techniques for large-scale PPI mapping are increasingly being applied to new organisms, including human, there is a corresponding need to rigorously evaluate, benchmark, and impartially filter the results. This chapter explores... Springer protocols feed by Bioinformatics news http://www.medworm.com/rss/comments.php?id=5179385 Wed, 31 Aug 2011 05:11:29 +0100 5179385 http://www.medworm.com/index.php?rid=5179384&cid=s_37118_79_f&fid=37118&url=http%3A%2F%2Fwww.springerprotocols.com%2FAbstract%2Fdoi%2F10.1007%2F978-1-61779-276-2_13 Protein–protein interactions are key to many aspects of the cell, including its cytoskeletal structure, the signaling processes in which it is involved, or its metabolism. Failure to form protein complexes or signaling cascades may sometimes translate into pathologic conditions such as cancer or neurodegenerative diseases. The set of all protein interactions between the proteins encoded by an organism constitutes its protein interaction network, representing a scaffold for biological function. Knowing the protein interaction network of an organism, combined with other sources of biological information, can unravel fundamental biological circuits and may help better understand the molecular basics of human diseases. The protein interaction network of an organism can be mapped by combi... Springer protocols feed by Bioinformatics news http://www.medworm.com/rss/comments.php?id=5179384 Wed, 31 Aug 2011 05:11:28 +0100 5179384 http://www.medworm.com/index.php?rid=5179383&cid=s_37118_79_f&fid=37118&url=http%3A%2F%2Fwww.springerprotocols.com%2FAbstract%2Fdoi%2F10.1007%2F978-1-61779-276-2_12 Gene-set enrichment analysis finds functionally coherent gene-sets, such as pathways, that are statistically overrepresented in a given gene list. Ideally, the number of resulting sets is smaller than the number of genes in the list, thus simplifying interpretation. However, the increasing number and redundancy of ­gene-sets used by many current enrichment analysis resources work against this ideal. “Enrichment Map” is a Cytoscape plug-in that helps overcome gene-set redundancy and aids in the interpretation of enrichment results. Gene-sets are organized in a network, where each set is a node and links represent gene overlap between sets. Automated network layout groups related gene-sets into ­network clusters, enabling the user to quickly identify the major enriched fu... Springer protocols feed by Bioinformatics news http://www.medworm.com/rss/comments.php?id=5179383 Wed, 31 Aug 2011 05:11:28 +0100 5179383 http://www.medworm.com/index.php?rid=5179382&cid=s_37118_79_f&fid=37118&url=http%3A%2F%2Fwww.springerprotocols.com%2FAbstract%2Fdoi%2F10.1007%2F978-1-61779-276-2_11 Bioinformatic methods to predict protein–protein interactions (PPI) via coevolutionary analysis have ­positioned themselves to compete alongside established in vitro methods, despite a lack of understanding for the underlying molecular mechanisms of the coevolutionary process. Investigating the alignment of coevolutionary predictions of PPI with experimental data can focus the effective scope of prediction and lead to better accuracies. A new rate-based coevolutionary method, MMM, preferentially finds obligate interacting proteins that form complexes, conforming to results from studies based on coimmunoprecipitation coupled with mass spectrometry. Using gold-standard databases as a benchmark for accuracy, MMM surpasses methods based on abundance ratios, suggesting that correlated... Springer protocols feed by Bioinformatics news http://www.medworm.com/rss/comments.php?id=5179382 Wed, 31 Aug 2011 05:11:27 +0100 5179382 http://www.medworm.com/index.php?rid=5179381&cid=s_37118_79_f&fid=37118&url=http%3A%2F%2Fwww.springerprotocols.com%2FAbstract%2Fdoi%2F10.1007%2F978-1-61779-276-2_10 DNA sequences are important sources of data for phylogenetic analysis. Nowadays, DNA sequencing is a routine technique in molecular biology laboratories. However, there are specific questions associated with project design and sequencing of plant samples for phylogenetic analysis, which may not be familiar to researchers starting in the field. This chapter gives an overview of methods and protocols involved in the sequencing of plant samples, including general recommendations on the selection of species/taxa and DNA regions to be sequenced, and field collection of plant samples. Protocols of plant sample preparation, DNA extraction, PCR and cloning, which are critical to the success of molecular phylogenetic projects, are described in detail. Common problems of sequencing (using the Sanger... Springer protocols feed by Bioinformatics news http://www.medworm.com/rss/comments.php?id=5179381 Wed, 31 Aug 2011 05:11:27 +0100 5179381 http://www.medworm.com/index.php?rid=4591118&cid=s_37118_79_f&fid=37118&url=http%3A%2F%2Fwww.springerprotocols.com%2FAbstract%2Fdoi%2F10.1007%2F978-1-61779-027-0_1 We provide an overview on the state of the art for the Omics technologies, the types of omics data and the bioinformatics resources relevant and related to Omics. We also illustrate the bioinformatics challenges of dealing with high-throughput data. This overview touches several fundamental aspects of Omics and bioinformatics: data standardisation, data sharing, storing Omics data appropriately and exploring Omics data in bioinformatics. Though the principles and concepts presented are true for the various different technological fields, we concentrate in three main Omics fields namely: genomics, transcriptomics and proteomics. Finally we address the integration of Omics data, and provide several useful links for bioinformatics and Omics. (Source: Springer protocols feed by Bioinformatics) Springer protocols feed by Bioinformatics news http://www.medworm.com/rss/comments.php?id=4591118 Tue, 15 Mar 2011 22:49:13 +0100 4591118 http://www.medworm.com/index.php?rid=4591117&cid=s_37118_79_f&fid=37118&url=http%3A%2F%2Fwww.springerprotocols.com%2FAbstract%2Fdoi%2F10.1007%2F978-1-61779-027-0_2 To facilitate sharing of Omics data, many groups of scientists have been working to establish the relevant data standards. The main components of data sharing standards are experiment description standards, data exchange standards, terminology standards, and experiment execution standards. Here we provide a survey of existing and emerging standards that are intended to assist the free and open exchange of large-format data. (Source: Springer protocols feed by Bioinformatics) Springer protocols feed by Bioinformatics news http://www.medworm.com/rss/comments.php?id=4591117 Tue, 15 Mar 2011 22:49:13 +0100 4591117 http://www.medworm.com/index.php?rid=4591116&cid=s_37118_79_f&fid=37118&url=http%3A%2F%2Fwww.springerprotocols.com%2FAbstract%2Fdoi%2F10.1007%2F978-1-61779-027-0_3 Technological Omics breakthroughs, including next generation sequencing, bring avalanches of data which need to undergo effective data management to ensure integrity, security, and maximal knowledge-gleaning. Data management system requirements include flexible input formats, diverse data entry mechanisms and views, user friendliness, attention to standards, hardware and software platform definition, as well as robustness. Relevant solutions elaborated by the scientific community include Laboratory Information Management Systems (LIMS) and standardization protocols facilitating data sharing and managing. In project planning, special consideration has to be made when choosing relevant Omics annotation sources, since many of them overlap and require sophisticated integration heuristics. The ... Springer protocols feed by Bioinformatics news http://www.medworm.com/rss/comments.php?id=4591116 Tue, 15 Mar 2011 22:49:13 +0100 4591116 http://www.medworm.com/index.php?rid=4591115&cid=s_37118_79_f&fid=37118&url=http%3A%2F%2Fwww.springerprotocols.com%2FAbstract%2Fdoi%2F10.1007%2F978-1-61779-027-0_4 Cross-Omics studies aimed at characterizing a specific phenotype on multiple levels are entering the ­scientific literature, and merging e.g. transcriptomics and proteomics data clearly promises to improve Omics data interpretation. Also for Systems Biology the integration of multi-level Omics profiles (also across species) is considered as central element. Due to the complexity of each specific Omics technique, specialization of experimental and bioinformatics research groups have become necessary, in turn demanding collaborative efforts for effectively implementing cross-Omics. This setting imposes specific emphasis on data sharing platforms for Omics data integration and cross-Omics data analysis and interpretation. Here we describe a software concept and methodology fostering Omics... Springer protocols feed by Bioinformatics news http://www.medworm.com/rss/comments.php?id=4591115 Tue, 15 Mar 2011 22:49:13 +0100 4591115 http://www.medworm.com/index.php?rid=4591114&cid=s_37118_79_f&fid=37118&url=http%3A%2F%2Fwww.springerprotocols.com%2FAbstract%2Fdoi%2F10.1007%2F978-1-61779-027-0_5 In Omics experiments, typically thousands of hypotheses are tested simultaneously, each based on very few independent replicates. Traditional tests like the t-test were shown to perform poorly with this new type of data. Furthermore, simultaneous consideration of many hypotheses, each prone to a decision error, requires powerful adjustments for this multiple testing situation. After a general introduction to statistical testing, we present the moderated t-statistic, the SAM statistic, and the RankProduct statistic which have been developed to evaluate hypotheses in typical Omics experiments. We also provide an introduction to the multiple testing problem and discuss some state-of-the-art procedures to address this issue. The presented test statistics are subjected to a comparative analysis... Springer protocols feed by Bioinformatics news http://www.medworm.com/rss/comments.php?id=4591114 Tue, 15 Mar 2011 22:49:13 +0100 4591114 http://www.medworm.com/index.php?rid=4591113&cid=s_37118_79_f&fid=37118&url=http%3A%2F%2Fwww.springerprotocols.com%2FAbstract%2Fdoi%2F10.1007%2F978-1-61779-027-0_6 Multiple Omics datasets (for example, high throughput mRNA and protein measurements for the same set of genes) are beginning to appear more widely within the fields of bioinformatics and computational biology. There are many tools available for the analysis of single datasets but two (or more) sets of coupled observations present more of a challenge. I describe some of the methods available – from classical statistical techniques to more recent advances from the fields of Machine Learning and Pattern Recognition for linking Omics data levels with particular focus on transcriptomics and proteomics profiles. (Source: Springer protocols feed by Bioinformatics) Springer protocols feed by Bioinformatics news http://www.medworm.com/rss/comments.php?id=4591113 Tue, 15 Mar 2011 22:49:13 +0100 4591113 http://www.medworm.com/index.php?rid=4591112&cid=s_37118_79_f&fid=37118&url=http%3A%2F%2Fwww.springerprotocols.com%2FAbstract%2Fdoi%2F10.1007%2F978-1-61779-027-0_7 Over the past 20 years, Omics technologies emerged as the consensual denomination of holistic molecular profiling. These techniques enable parallel measurements of biological -omes, or “all constituents considered collectively”, and utilize the latest advancements in transcriptomics, proteomics, metabolomics, imaging, and bioinformatics. The technological accomplishments in increasing the sensitivity and throughput of the analytical devices, the standardization of the protocols and the widespread availability of reagents made the capturing of static molecular portraits of biological systems a routine task. The next generation of time course molecular profiling already allows for extensive molecular snapshots to be taken along the trajectory of time evolution of the investigated... Springer protocols feed by Bioinformatics news http://www.medworm.com/rss/comments.php?id=4591112 Tue, 15 Mar 2011 22:49:13 +0100 4591112 http://www.medworm.com/index.php?rid=4591111&cid=s_37118_79_f&fid=37118&url=http%3A%2F%2Fwww.springerprotocols.com%2FAbstract%2Fdoi%2F10.1007%2F978-1-61779-027-0_8 The diverse fields of Omics research share a common logical structure combining a cataloging effort for a particular class of molecules or interactions, the underlying -ome, and a quantitative aspect attempting to record spatiotemporal patterns of concentration, expression, or variation. Consequently, these fields also share a common set of difficulties and limitations. In spite of the great success stories of Omics projects over the last decade, much remains to be understood not only at the technological, but also at the conceptual level. Here, we focus on the dark corners of Omics research, where the problems, limitations, conceptual difficulties, and lack of knowledge are hidden. (Source: Springer protocols feed by Bioinformatics) Springer protocols feed by Bioinformatics news http://www.medworm.com/rss/comments.php?id=4591111 Tue, 15 Mar 2011 22:49:13 +0100 4591111 http://www.medworm.com/index.php?rid=4591110&cid=s_37118_79_f&fid=37118&url=http%3A%2F%2Fwww.springerprotocols.com%2FAbstract%2Fdoi%2F10.1007%2F978-1-61779-027-0_10 We describe and discuss methods used to identify SNPs associated with disease in case–control studies. An outline on study population selection, sample collection and genotyping platforms is presented, complemented by SNP selection, data preprocessing and analysis. (Source: Springer protocols feed by Bioinformatics) Springer protocols feed by Bioinformatics news http://www.medworm.com/rss/comments.php?id=4591110 Tue, 15 Mar 2011 22:49:13 +0100 4591110 http://www.medworm.com/index.php?rid=4591109&cid=s_37118_79_f&fid=37118&url=http%3A%2F%2Fwww.springerprotocols.com%2FAbstract%2Fdoi%2F10.1007%2F978-1-61779-027-0_11 Copy number variation is known to be an important component of structural variation in the human genome. Greater than 1 kb in size, these gains and losses of genetic material are known to confer risk to many human diseases, both Mendelian and complex. Therefore, the technologies used to detect copy number variation have been quickly improving in both throughput and cost. From comparative genomic hybridization to synthetic high-density oligonucleotide arrays to next-generation sequencing methods, algorithms used to estimate copy number are plentiful. Here we describe a practical introduction to the copy number variation technology and available analysis methods, and demonstrate the analysis flow on an example case. (Source: Springer protocols feed by Bioinformatics) Springer protocols feed by Bioinformatics news http://www.medworm.com/rss/comments.php?id=4591109 Tue, 15 Mar 2011 22:49:13 +0100 4591109 http://www.medworm.com/index.php?rid=4591108&cid=s_37118_79_f&fid=37118&url=http%3A%2F%2Fwww.springerprotocols.com%2FAbstract%2Fdoi%2F10.1007%2F978-1-61779-027-0_13 Transcriptomics has played an essential role as proof of concept in the development of experimental and bioinformatics approaches for the generation and analysis of Omics data. We are giving an introduction on how large-scale technologies for gene expression profiling, especially microarrays, have changed the view from studying single molecular events to a systems level view of global mechanisms in a cell, the biological processes, and their pathological mutations. The main platforms available for gene expression profiling (from microarrays to RNA-seq) are presented and the general concepts that need to be taken into account for proper data analysis in order to extract objective and general conclusions from transcriptomics experiments are introduced. We also describe the available main bio... Springer protocols feed by Bioinformatics news http://www.medworm.com/rss/comments.php?id=4591108 Tue, 15 Mar 2011 22:49:13 +0100 4591108 http://www.medworm.com/index.php?rid=4591107&cid=s_37118_79_f&fid=37118&url=http%3A%2F%2Fwww.springerprotocols.com%2FAbstract%2Fdoi%2F10.1007%2F978-1-61779-027-0_14 We describe current approaches to identify genome-wide functional RNA transcripts (experimentally as well as computationally), and focus on computational methods that may be utilized to disclose their function. While genome databases offer a wealth of information about known and putative functions for protein-coding genes, functional information for novel non-coding RNA genes is almost nonexistent. This is mainly explained by the lack of established software tools to efficiently reveal the function and evolutionary origin of non-coding RNA genes. Here, we describe in detail computational approaches one may follow to annotate and classify an RNA transcript. (Source: Springer protocols feed by Bioinformatics) Springer protocols feed by Bioinformatics news http://www.medworm.com/rss/comments.php?id=4591107 Tue, 15 Mar 2011 22:49:13 +0100 4591107 http://www.medworm.com/index.php?rid=4591106&cid=s_37118_79_f&fid=37118&url=http%3A%2F%2Fwww.springerprotocols.com%2FAbstract%2Fdoi%2F10.1007%2F978-1-61779-027-0_15 This article addresses the key algorithmic problems bioinformaticians face when handling modern proteomic samples and shows common solutions to them. We provide examples on how algorithms can be combined to build relatively complex analysis pipelines, point out certain pitfalls and aspects worth considering and give a list of current state-of-the-art tools. (Source: Springer protocols feed by Bioinformatics) Springer protocols feed by Bioinformatics news http://www.medworm.com/rss/comments.php?id=4591106 Tue, 15 Mar 2011 22:49:13 +0100 4591106 http://www.medworm.com/index.php?rid=4591105&cid=s_37118_79_f&fid=37118&url=http%3A%2F%2Fwww.springerprotocols.com%2FAbstract%2Fdoi%2F10.1007%2F978-1-61779-027-0_16 The broad view of the state of biological systems cannot be complete without the added value of integrating proteomic and genomic data with metabolite measurement. By definition, metabolomics aims at quantifying not less than the totality of small molecules present in a biofluid, tissue, organism, or any material beyond living systems. To cope with the complexity of the task, mass spectrometry (MS) is the most promising analytical environment to fulfill increasing appetite for more accurate and larger view of the metabolome while providing sufficient data generation throughput. Bioinformatics and associated disciplines naturally play a central role in bridging the gap between fast evolving technology and domain experts. Here, we describe the strategies to translate crude MS information int... Springer protocols feed by Bioinformatics news http://www.medworm.com/rss/comments.php?id=4591105 Tue, 15 Mar 2011 22:49:13 +0100 4591105 http://www.medworm.com/index.php?rid=4591104&cid=s_37118_79_f&fid=37118&url=http%3A%2F%2Fwww.springerprotocols.com%2FAbstract%2Fdoi%2F10.1007%2F978-1-61779-027-0_9 The advent of High Throughput Sequencing (HTS) methods opens new opportunities for the analysis of genomes and transcriptomes. While the sequencing of a whole mammalian genome took several years at the turn of this century, today it is only a matter of weeks. The race towards the thousand-dollar genome is fueled by the – ethically challenging – idea of personalized genomic medicine. However, these methods allow new and interesting insights in many aspects such as the discovery of novel noncoding RNA classes, structural variants, or alternative splice sites to name a few. Meanwhile, several methods for HTS have been introduced to the markets. Here, an overview on the technologies and the bioinformatics analysis of HTS data is given. (Source: Springer protocols feed by Bioinforma... Springer protocols feed by Bioinformatics news http://www.medworm.com/rss/comments.php?id=4591104 Tue, 15 Mar 2011 22:49:13 +0100 4591104 http://www.medworm.com/index.php?rid=4591103&cid=s_37118_79_f&fid=37118&url=http%3A%2F%2Fwww.springerprotocols.com%2FAbstract%2Fdoi%2F10.1007%2F978-1-61779-027-0_17 Progress in experimental procedures has led to rapid availability of Omics profiles. Various open-access as well as commercial tools have been developed for storage, analysis, and interpretation of transcriptomics, proteomics, and metabolomics data. Generally, major analysis steps include data storage, retrieval, preprocessing, and normalization, followed by identification of differentially expressed features, functional annotation on the level of biological processes and molecular pathways, as well as interpretation of gene lists in the context of protein–protein interaction networks. In this chapter, we discuss a sequential transcriptomics data analysis workflow utilizing open-source tools, specifically exemplified on a gene expression dataset on familial hypercholesterolemia. (Sou... Springer protocols feed by Bioinformatics news http://www.medworm.com/rss/comments.php?id=4591103 Tue, 15 Mar 2011 22:49:13 +0100 4591103 http://www.medworm.com/index.php?rid=4591102&cid=s_37118_79_f&fid=37118&url=http%3A%2F%2Fwww.springerprotocols.com%2FAbstract%2Fdoi%2F10.1007%2F978-1-61779-027-0_18 “-Omics” is a current suffix for numerous types of large-scale biological data generation procedures, which naturally demand the development of novel algorithms for data storage and analysis. With next generation genome sequencing burgeoning, it is pivotal to decipher a coding site on the genome, a gene’s function, and information on transcripts next to the pure availability of sequence information. To explore a genome and downstream molecular processes, we need umpteen results at the various levels of cellular organization by utilizing different experimental designs, data analysis strategies and methodologies. Here comes the need for controlled vocabularies and data integration to annotate, store, and update the flow of experimental data. This chapter explores key method... Springer protocols feed by Bioinformatics news http://www.medworm.com/rss/comments.php?id=4591102 Tue, 15 Mar 2011 22:49:12 +0100 4591102 http://www.medworm.com/index.php?rid=4591101&cid=s_37118_79_f&fid=37118&url=http%3A%2F%2Fwww.springerprotocols.com%2FAbstract%2Fdoi%2F10.1007%2F978-1-61779-027-0_19 Omics data and computational approaches are today providing a key to disentangle the complex architecture of living systems. The integration and analysis of data of different nature allows to extract meaningful representations of signaling pathways and protein interactions networks, helpful in achieving an increased understanding of such intricate biochemical processes. We here describe a general workflow and relative hurdles in integrating online Omics data and analyzing reconstructed representations by using the available computational platforms. (Source: Springer protocols feed by Bioinformatics) Springer protocols feed by Bioinformatics news http://www.medworm.com/rss/comments.php?id=4591101 Tue, 15 Mar 2011 22:49:12 +0100 4591101 http://www.medworm.com/index.php?rid=4591100&cid=s_37118_79_f&fid=37118&url=http%3A%2F%2Fwww.springerprotocols.com%2FAbstract%2Fdoi%2F10.1007%2F978-1-61779-027-0_20 We provide a commented overview of the available databases for the systematic collection of pathway information and biological models essential for the interpretation of Omics data. Then, we present both the state of the art and the future challenges of network inference, a research area dealing with the deduction of reaction mechanisms from experimental Omics data. This approach represents one of the most challenging instances for making use of the huge amount of information gathered in the Omics era. (Source: Springer protocols feed by Bioinformatics) Springer protocols feed by Bioinformatics news http://www.medworm.com/rss/comments.php?id=4591100 Tue, 15 Mar 2011 22:49:12 +0100 4591100 http://www.medworm.com/index.php?rid=4591099&cid=s_37118_79_f&fid=37118&url=http%3A%2F%2Fwww.springerprotocols.com%2FAbstract%2Fdoi%2F10.1007%2F978-1-61779-027-0_21 We describe approaches to identify biological concepts in the form of Medical Subject Headings (MeSH terms) as extracted from MEDLINE that are significantly overrepresented within the identified gene set relative to those associated with the overall collection of genes on the underlying Omics platform. The method’s principle strength is its ability to simultaneously depict similarities that may exist at the level of biological structure, molecular function, physiology, genetics, and clinically manifest diseases, just as a single published article about a gene of interest may report findings within several of these same dimensions. (Source: Springer protocols feed by Bioinformatics) Springer protocols feed by Bioinformatics news http://www.medworm.com/rss/comments.php?id=4591099 Tue, 15 Mar 2011 22:49:12 +0100 4591099 http://www.medworm.com/index.php?rid=4591098&cid=s_37118_79_f&fid=37118&url=http%3A%2F%2Fwww.springerprotocols.com%2FAbstract%2Fdoi%2F10.1007%2F978-1-61779-027-0_22 The Omics revolution has provided the researcher with tools and methodologies for qualitative and quantitative assessment of a wide spectrum of molecular players spanning from the genome to the meta­bolome level. As a consequence, explorative analysis (in contrast to purely hypothesis driven research procedures) has become applicable. However, numerous issues have to be considered for deriving meaningful results from Omics, and bioinformatics has to respect these in data analysis and interpretation. Aspects include sample type and quality, concise definition of the (clinical) question, and selection of samples ideally coming from thoroughly defined sample and data repositories. Omics suffers from a principal shortcoming, namely unbalanced sample-to-feature matrix denoted as “curs... Springer protocols feed by Bioinformatics news http://www.medworm.com/rss/comments.php?id=4591098 Tue, 15 Mar 2011 22:49:12 +0100 4591098 http://www.medworm.com/index.php?rid=4591097&cid=s_37118_79_f&fid=37118&url=http%3A%2F%2Fwww.springerprotocols.com%2FAbstract%2Fdoi%2F10.1007%2F978-1-61779-027-0_23 Owing to the growing knowledge about the cellular molecular network and its alterations in diseases, most of the diseases become considered as “systems distortion of the cellular molecular network”. This view of diseases, which we call “systems pathology”, has brought about a new usage of the disease Omics, that is, to identify the altered molecular network underlying the disease. In this chapter, we discuss the technologies and clinical applications for Omics-based identification of pathophysiological process. In doing so, we classify the methods into two classes: one is a “data-inductive approach” which infers gene regulatory and transcriptional networks by gene expression data from DNA microarrays, and the other is a “knowledge-referenced approa... Springer protocols feed by Bioinformatics news http://www.medworm.com/rss/comments.php?id=4591097 Tue, 15 Mar 2011 22:49:12 +0100 4591097 http://www.medworm.com/index.php?rid=4591096&cid=s_37118_79_f&fid=37118&url=http%3A%2F%2Fwww.springerprotocols.com%2FAbstract%2Fdoi%2F10.1007%2F978-1-61779-027-0_24 We present an overview of general data mining methods and their applications to biomarker discovery with particular focus on genomics and proteomics data. Two case studies are exemplarily presented, and relevant data mining terminology and techniques are explained. (Source: Springer protocols feed by Bioinformatics) Springer protocols feed by Bioinformatics news http://www.medworm.com/rss/comments.php?id=4591096 Tue, 15 Mar 2011 22:49:12 +0100 4591096 http://www.medworm.com/index.php?rid=4591095&cid=s_37118_79_f&fid=37118&url=http%3A%2F%2Fwww.springerprotocols.com%2FAbstract%2Fdoi%2F10.1007%2F978-1-61779-027-0_25 The exponential growth of high-throughput Omics data has provided an unprecedented opportunity for new target identification to fuel the dried-up drug discovery pipeline. However, the bioinformatics analysis of large amount and heterogeneous Omics data has posed a great deal of technical challenges for experimentalists who lack statistical skills. Moreover, due to the complexity of human diseases, it is essential to analyze the Omics data in the context of molecular networks to detect meaningful biological targets and understand disease processes. Here, we describe an integrated bioinformatics analysis strategy and provide a running example to identify suitable targets for our in-house Enzyme-Mediated Cancer Imaging and Therapy (EMCIT) technology. In addition, we go through a few key conce... Springer protocols feed by Bioinformatics news http://www.medworm.com/rss/comments.php?id=4591095 Tue, 15 Mar 2011 22:49:12 +0100 4591095 http://www.medworm.com/index.php?rid=4591094&cid=s_37118_79_f&fid=37118&url=http%3A%2F%2Fwww.springerprotocols.com%2FAbstract%2Fdoi%2F10.1007%2F978-1-61779-027-0_26 We describe a downstream workflow and procedures for functional analysis that focus on biological pathways, from which molecular targets can be derived and proposed for experimental validation. (Source: Springer protocols feed by Bioinformatics) Springer protocols feed by Bioinformatics news http://www.medworm.com/rss/comments.php?id=4591094 Tue, 15 Mar 2011 22:49:12 +0100 4591094 http://www.medworm.com/index.php?rid=4060047&cid=s_37118_79_f&fid=37118&url=http%3A%2F%2Fwww.springerprotocols.com%2FAbstract%2Fdoi%2F10.1007%2F978-1-60761-580-4_23 This chapter is an introductory reference guide highlighting some of the most common statistical topics, broken down into both command-line syntax and graphical interface point-and-click commands. This chapter serves to supplement more formal statistics lessons and expedite using Stata to compute basic analyses. (Source: Springer protocols feed by Bioinformatics) Springer protocols feed by Bioinformatics news http://www.medworm.com/rss/comments.php?id=4060047 Mon, 25 Jan 2010 00:00:00 +0100 4060047 http://www.medworm.com/index.php?rid=4060046&cid=s_37118_79_f&fid=37118&url=http%3A%2F%2Fwww.springerprotocols.com%2FAbstract%2Fdoi%2F10.1007%2F978-1-60761-580-4_22 A majority of original articles published in biomedical journals include some form of statistical analysis. Unfortunately, many of the articles contain errors in statistical design and/or analysis. These errors are worrisome, as the misuse of statistics jeopardizes the process of scientific discovery and the accumulation of scientific knowledge. To help avoid these errors and improve statistical reporting, four approaches are suggested: (1) development of guidelines for statistical reporting that could be adopted by all journals, (2) improvement in statistics curricula in biomedical research programs with an emphasis on hands-on teaching by biostatisticians, (3) expansion and enhancement of biomedical science curricula in statistics programs, and (4) increased participation of biostatistic... Springer protocols feed by Bioinformatics news http://www.medworm.com/rss/comments.php?id=4060046 Mon, 25 Jan 2010 00:00:00 +0100 4060046 http://www.medworm.com/index.php?rid=4060045&cid=s_37118_79_f&fid=37118&url=http%3A%2F%2Fwww.springerprotocols.com%2FAbstract%2Fdoi%2F10.1007%2F978-1-60761-580-4_21 Cardiovascular disease, metabolic syndrome, schizophrenia, diabetes, bipolar disorder, and autism are a few of the numerous complex diseases for which researchers are trying to decipher the genetic composition. One interest of geneticists is to determine the quantitative trait loci (QTLs) that underlie the genetic portion of these diseases and their risk factors. The difficulty for researchers is that the QTLs underlying these diseases are likely to have small to medium effects which will necessitate having large studies in order to have adequate power. Combining information across multiple studies provides a way for researchers to potentially increase power while making the most of existing studies. (Source: Springer protocols feed by Bioinformatics) Springer protocols feed by Bioinformatics news http://www.medworm.com/rss/comments.php?id=4060045 Mon, 25 Jan 2010 00:00:00 +0100 4060045 http://www.medworm.com/index.php?rid=4060044&cid=s_37118_79_f&fid=37118&url=http%3A%2F%2Fwww.springerprotocols.com%2FAbstract%2Fdoi%2F10.1007%2F978-1-60761-580-4_20 Many biomedical applications are concerned with the problem of selecting important predictors from a high-dimensional set of candidates, with the gene expression data as one example. Due to the fact that the sample size in any single study is usually small, it is thus important to combine information from multiple studies. In this chapter, we introduce a Bayesian hierarchical modeling approach which models study-to-study heterogeneity explicitly to borrow strength across studies. Using a carefully formulated prior specification, we develop a fast approach to predictor selection and shrinkage estimation for high-dimensional predictors. The proposed approach, which is related to the relevance vector machine (RVM), relies on maximum a posteriori (MAP) estimation to rapidly obtain a sparse est... Springer protocols feed by Bioinformatics news http://www.medworm.com/rss/comments.php?id=4060044 Mon, 25 Jan 2010 00:00:00 +0100 4060044 http://www.medworm.com/index.php?rid=4060043&cid=s_37118_79_f&fid=37118&url=http%3A%2F%2Fwww.springerprotocols.com%2FAbstract%2Fdoi%2F10.1007%2F978-1-60761-580-4_19 Large-scale sequencing, copy number, mRNA, and protein data have given great promise to the biomedical research, while posing great challenges to data management and data analysis. Integrating different types of high-throughput data from diverse sources can increase the statistical power of data analysis and provide deeper biological understanding. This chapter uses two biomedical research examples to illustrate why there is an urgent need to develop reliable and robust methods for integrating the heterogeneous data. We then introduce and review some recently developed statistical methods for integrative analysis for both statistical inference and classification purposes. Finally, we present some useful public access databases and program code to facilitate the integrative analysis in prac... Springer protocols feed by Bioinformatics news http://www.medworm.com/rss/comments.php?id=4060043 Mon, 25 Jan 2010 00:00:00 +0100 4060043 http://www.medworm.com/index.php?rid=4060042&cid=s_37118_79_f&fid=37118&url=http%3A%2F%2Fwww.springerprotocols.com%2FAbstract%2Fdoi%2F10.1007%2F978-1-60761-580-4_18 During the last decade, analytical methods for the detection and quantification of proteins and peptides in biological samples have been considerably improved. It is therefore now possible to compare simultaneously the expression levels of hundreds or thousands of proteins in different types of tissue, for example, normal and cancerous, or in different cell lines. In this chapter, we illustrate statistical designs for such proteomics experiments as well as methods for the analysis of resulting data. In particular, we focus on the preprocessing and analysis of protein expression levels recorded by the use of either two-dimensional gel electrophoresis or mass spectrometry. (Source: Springer protocols feed by Bioinformatics) Springer protocols feed by Bioinformatics news http://www.medworm.com/rss/comments.php?id=4060042 Mon, 25 Jan 2010 00:00:00 +0100 4060042 http://www.medworm.com/index.php?rid=4060041&cid=s_37118_79_f&fid=37118&url=http%3A%2F%2Fwww.springerprotocols.com%2FAbstract%2Fdoi%2F10.1007%2F978-1-60761-580-4_17 The analysis of genome-wide association studies (GWAS) poses statistical hurdles that have to be handled efficiently in order for the study to be successful. The two largest impediments in the analysis phase of the study are the multiple comparisons problem and maintaining robustness against confounding due to population admixture and stratification. For quantitative traits in family-based designs, Van Steen (1) proposed a two-stage testing strategy that can be considered a hybrid approach between family-based and population-based analysis. By including the population-based component into the family-based analysis, the Van Steen algorithm maximizes the statistical power, while at the same time, maintains the original robustness of family-based association tests (FBATs) (2–4). The Van... Springer protocols feed by Bioinformatics news http://www.medworm.com/rss/comments.php?id=4060041 Mon, 25 Jan 2010 00:00:00 +0100 4060041 http://www.medworm.com/index.php?rid=4060040&cid=s_37118_79_f&fid=37118&url=http%3A%2F%2Fwww.springerprotocols.com%2FAbstract%2Fdoi%2F10.1007%2F978-1-60761-580-4_16 Gene expression profiling technique now enables scientists to obtain a genome-wide picture of cellular functions on various human disease mechanisms which has also proven to be extremely valuable in forecasting patients’ prognosis and therapeutic responses. A wide range of multivariate techniques have been employed in biomedical applications on such expression profiling data in order to identify expression biomarkers that are highly associated with patients’ clinical outcome and to train multi-gene prediction models that can forecast various human disease outcome and drug toxicities. We provide here a brief overview on some of these approaches, succinctly summarizing relevant basic concepts, statistical algorithms, and several practical applications. We also introduce our recen... Springer protocols feed by Bioinformatics news http://www.medworm.com/rss/comments.php?id=4060040 Mon, 25 Jan 2010 00:00:00 +0100 4060040 http://www.medworm.com/index.php?rid=4060039&cid=s_37118_79_f&fid=37118&url=http%3A%2F%2Fwww.springerprotocols.com%2FAbstract%2Fdoi%2F10.1007%2F978-1-60761-580-4_15 High-throughput technologies can routinely assay biological or clinical samples and produce wide data sets where each sample is associated with tens of thousands of measurements. Such data sets can be mined to discover biomarkers and develop statistical models capable of predicting an endpoint of interest from data measured in the samples. The field of biomarker model development combines methods from statistics and machine learning to develop and evaluate predictive biomarker models. In this chapter, we discuss the computational steps involved in the development of biomarker models designed to predict information about individual samples and review approaches often used to implement each step. A practical example of biomarker model development in a large gene expression data set is presen... Springer protocols feed by Bioinformatics news http://www.medworm.com/rss/comments.php?id=4060039 Mon, 25 Jan 2010 00:00:00 +0100 4060039 http://www.medworm.com/index.php?rid=4060038&cid=s_37118_79_f&fid=37118&url=http%3A%2F%2Fwww.springerprotocols.com%2FAbstract%2Fdoi%2F10.1007%2F978-1-60761-580-4_14 With advancing of modern technologies, high-dimensional data have prevailed in computational biology. The number of variables p is very large, and in many applications, p is larger than the number of observational units n. Such high dimensionality and the unconventional small-n-large-p setting have posed new challenges to statistical analysis methods. Dimension reduction, which aims to reduce the predictor dimension prior to any modeling efforts, offers a potentially useful avenue to tackle such high-dimensional regression. In this chapter, we review a number of commonly used dimension reduction approaches, including principal component analysis, partial least squares, and sliced inverse regression. For each method, we review its background and its applications in computational biology, di... Springer protocols feed by Bioinformatics news http://www.medworm.com/rss/comments.php?id=4060038 Mon, 25 Jan 2010 00:00:00 +0100 4060038 http://www.medworm.com/index.php?rid=4060037&cid=s_37118_79_f&fid=37118&url=http%3A%2F%2Fwww.springerprotocols.com%2FAbstract%2Fdoi%2F10.1007%2F978-1-60761-580-4_13 Hidden Markov models have wide applications in pattern recognition. In genome sequence analysis, hidden Markov models (HMMs) have been applied to the identification of regions of the genome that contain regulatory information, i.e., binding sites. In higher eukaryotes, the regulatory information is organized into modular units called cis-regulatory modules. Each module contains multiple binding sites for a specific combination of several transcription factors. In this chapter, we gave a brief review of hidden Markov models, standard algorithms from HMM, and their applications to motif findings. We then introduce the application of HMM to a complex system in which an HMM is combined with Bayesian inference to identify transcription factor binding sites and cis-regulatory modules. (Source: S... Springer protocols feed by Bioinformatics news http://www.medworm.com/rss/comments.php?id=4060037 Mon, 25 Jan 2010 00:00:00 +0100 4060037 http://www.medworm.com/index.php?rid=4060036&cid=s_37118_79_f&fid=37118&url=http%3A%2F%2Fwww.springerprotocols.com%2FAbstract%2Fdoi%2F10.1007%2F978-1-60761-580-4_12 In molecular biology, we are often interested in determining the group structure in, e.g., a population of cells or microarray gene expression data. Clustering methods identify groups of similar observations, but the results can depend on the chosen method’s assumptions and starting parameter values. In this chapter, we give a broad overview of both attribute- and similarity-based clustering, describing both the methods and their performance. The parametric and nonparametric approaches presented vary in whether or not they require knowing the number of clusters in advance as well as the shapes of the estimated clusters. Additionally, we include a biclustering algorithm that incorporates variable selection into the clustering procedure. We finish with a discussion of some common metho... Springer protocols feed by Bioinformatics news http://www.medworm.com/rss/comments.php?id=4060036 Mon, 25 Jan 2010 00:00:00 +0100 4060036 http://www.medworm.com/index.php?rid=4060035&cid=s_37118_79_f&fid=37118&url=http%3A%2F%2Fwww.springerprotocols.com%2FAbstract%2Fdoi%2F10.1007%2F978-1-60761-580-4_11 The support vector machine is a supervised learning technique for classification increasingly used in many applications of data mining, engineering, and bioinformatics. This chapter aims to provide an introduction to the method, covering from the basic concept of the optimal separating hyperplane to its nonlinear generalization through kernels. A general framework of kernel methods that encompass the support vector machine as a special case is outlined. In addition, statistical properties that illuminate both advantage and limitation of the method due to its specific mechanism for classification are briefly discussed. For illustration of the method and related practical issues, an application to real data with high-dimensional features is presented. (Source: Springer protocols feed by Bioi... Springer protocols feed by Bioinformatics news http://www.medworm.com/rss/comments.php?id=4060035 Mon, 25 Jan 2010 00:00:00 +0100 4060035 http://www.medworm.com/index.php?rid=4060034&cid=s_37118_79_f&fid=37118&url=http%3A%2F%2Fwww.springerprotocols.com%2FAbstract%2Fdoi%2F10.1007%2F978-1-60761-580-4_9 Microarray experiments have become routine in the past few years in many fields of biology. Analysis of array hybridizations is often performed with the help of commercial software programs, which produce gene lists, graphs, and sometimes provide values for the statistical significance of the results. Exactly what is computed by many of the available programs is often not easy to reconstruct or may even be impossible to know for the end user. It is therefore not surprising that many biology students and some researchers using microarray data do not fully understand the nature of the underlying statistics used to arrive at the results. (Source: Springer protocols feed by Bioinformatics) Springer protocols feed by Bioinformatics news http://www.medworm.com/rss/comments.php?id=4060034 Mon, 25 Jan 2010 00:00:00 +0100 4060034 http://www.medworm.com/index.php?rid=4060033&cid=s_37118_79_f&fid=37118&url=http%3A%2F%2Fwww.springerprotocols.com%2FAbstract%2Fdoi%2F10.1007%2F978-1-60761-580-4_10 This chapter describes methods for learning gene interaction networks from high-throughput gene expression data sets. Many genes have unknown or poorly understood functions and interactions, especially in diseases such as cancer where the genome is frequently mutated. The gene interactions inferred by learning a network model from the data can form the basis of hypotheses that can be verified by subsequent biological experiments. This chapter focuses specifically on Bayesian network models, which have a level of mathematical detail greater than purely conceptual models but less than detailed differential equation models. From a network learning perspective the most severe problem with microarray data is the limited sample size, since there are usually many plausible networks for modeling t... Springer protocols feed by Bioinformatics news http://www.medworm.com/rss/comments.php?id=4060033 Mon, 25 Jan 2010 00:00:00 +0100 4060033 http://www.medworm.com/index.php?rid=4060032&cid=s_37118_79_f&fid=37118&url=http%3A%2F%2Fwww.springerprotocols.com%2FAbstract%2Fdoi%2F10.1007%2F978-1-60761-580-4_8 The rapid advances in biotechnology have given rise to a variety of high-dimensional data. Many of these data, including DNA microarray data, mass spectrometry protein data, and high-throughput screening (HTS) assay data, are generated by complex experimental procedures that involve multiple steps such as sample extraction, purification and/or amplification, labeling, fragmentation, and detection. Therefore, the quantity of interest is not directly obtained and a number of preprocessing procedures are necessary to convert the raw data into the format with biological relevance. This also makes exploratory data analysis and visualization essential steps to detect possible defects, anomalies or distortion of the data, to test underlying assumptions and thus ensure data quality. The characteri... Springer protocols feed by Bioinformatics news http://www.medworm.com/rss/comments.php?id=4060032 Mon, 25 Jan 2010 00:00:00 +0100 4060032 http://www.medworm.com/index.php?rid=4060031&cid=s_37118_79_f&fid=37118&url=http%3A%2F%2Fwww.springerprotocols.com%2FAbstract%2Fdoi%2F10.1007%2F978-1-60761-580-4_7 Epigenetics is the study of heritable change other than those encoded in DNA sequence. Cytosine methylation of DNA at CpG dinucleotides is the most well-studied epigenetic phenomenon, although epigenetic changes also encompass non-DNA methylation mechanisms, such as covalent histone modifications, micro-RNA interactions, and chromatin remodeling complexes. Methylation changes, both global and gene specific, have been observed to be associated with disease, particularly in cancer. (Source: Springer protocols feed by Bioinformatics) Springer protocols feed by Bioinformatics news http://www.medworm.com/rss/comments.php?id=4060031 Mon, 25 Jan 2010 00:00:00 +0100 4060031 http://www.medworm.com/index.php?rid=4060030&cid=s_37118_79_f&fid=37118&url=http%3A%2F%2Fwww.springerprotocols.com%2FAbstract%2Fdoi%2F10.1007%2F978-1-60761-580-4_6 Genetic linkage analysis has been a traditional means for identifying regions of the genome with large genetic effects that contribute to a disease. Following linkage analysis, association studies are widely pursued to fine-tune regions with significant linkage signals. For complex diseases which often involve function of multi-genetic variants each with small or moderate effect, linkage analysis has little power compared to association studies. In this chapter, we give a brief review of design issues related to linkage analysis and association studies with human genetic data. We introduce methods commonly used for linkage and association studies and compared the relative merits of the family-based and population-based association studies. Compared to candidate gene studies, a genomewide b... Springer protocols feed by Bioinformatics news http://www.medworm.com/rss/comments.php?id=4060030 Mon, 25 Jan 2010 00:00:00 +0100 4060030 http://www.medworm.com/index.php?rid=4060029&cid=s_37118_79_f&fid=37118&url=http%3A%2F%2Fwww.springerprotocols.com%2FAbstract%2Fdoi%2F10.1007%2F978-1-60761-580-4_5 Sample size calculation is a critical procedure when designing a new biological study. In this chapter, we consider molecular biology studies generating huge dimensional data. Microarray studies are typical examples, so that we state this chapter in terms of gene microarray data, but the discussed methods can be used for design and analysis of any molecular biology studies involving high-dimensional data. In this chapter, we discuss sample size calculation methods for molecular biology studies when the discovery of prognostic molecular markers is performed by accurately controlling false discovery rate (FDR) or family-wise error rate (FWER) in the final data analysis. We limit our discussion to the two-sample case. (Source: Springer protocols feed by Bioinformatics) Springer protocols feed by Bioinformatics news http://www.medworm.com/rss/comments.php?id=4060029 Mon, 25 Jan 2010 00:00:00 +0100 4060029 http://www.medworm.com/index.php?rid=4060028&cid=s_37118_79_f&fid=37118&url=http%3A%2F%2Fwww.springerprotocols.com%2FAbstract%2Fdoi%2F10.1007%2F978-1-60761-580-4_4 In this chapter we will explore Bayesian alternatives to the t-test. We saw in Chapter 1 how t-test can be used to test whether the expected outcomes of the two groups are equal or not. In Chapter 3 we saw how to make inferences from a Bayesian perspective in principle. In this chapter we will put these together to develop a Bayesian procedure for a t-test. This procedure depends on the data only through the t-statistic. It requires prior inputs and we will discuss how to assign them. We will use an example from a microarray study as to demonstrate the practical issues. The microarray study is an important application for the Bayesian t-test as it naturally brings up the question of simultaneous t-tests. It turns out that the Bayesian procedure can easily be extended to carry several t-tes... Springer protocols feed by Bioinformatics news http://www.medworm.com/rss/comments.php?id=4060028 Mon, 25 Jan 2010 00:00:00 +0100 4060028 http://www.medworm.com/index.php?rid=4060027&cid=s_37118_79_f&fid=37118&url=http%3A%2F%2Fwww.springerprotocols.com%2FAbstract%2Fdoi%2F10.1007%2F978-1-60761-580-4_3 Bayesian methods are rapidly becoming popular tools for making statistical inference in various fields of science including biology, engineering, finance, and genetics. One of the key aspects of Bayesian inferential method is its logical foundation that provides a coherent framework to utilize not only empirical but also scientific information available to a researcher. Prior knowledge arising from scientific background, expert judgment, or previously collected data is used to build a prior distribution which is then combined with current data via the likelihood function to characterize the current state of knowledge using the so-called posterior distribution. Bayesian methods allow the use of models of complex physical phenomena that were previously too difficult to estimate (e.g., using ... Springer protocols feed by Bioinformatics news http://www.medworm.com/rss/comments.php?id=4060027 Mon, 25 Jan 2010 00:00:00 +0100 4060027 http://www.medworm.com/index.php?rid=4060026&cid=s_37118_79_f&fid=37118&url=http%3A%2F%2Fwww.springerprotocols.com%2FAbstract%2Fdoi%2F10.1007%2F978-1-60761-580-4_2 In 2003, the completion of the Human Genome Project (1) together with advances in computational resources (2) were expected to launch an era where the genetic and genomic contributions to many common diseases would be found. In the years following, however, researchers became increasingly frustrated as most reported ‘findings’ could not be replicated in independent studies (3). To improve the signal/noise ratio, it was suggested to increase the number of cases to be included to tens of thousands (4), a requirement that would dramatically restrict the scope of personalized medicine. Similarly, there was little success in elucidating the gene–gene interactions involved in complex diseases or even in developing criteria for assessing their phenotypes. As a partial solution t... Springer protocols feed by Bioinformatics news http://www.medworm.com/rss/comments.php?id=4060026 Mon, 25 Jan 2010 00:00:00 +0100 4060026 http://www.medworm.com/index.php?rid=4060025&cid=s_37118_79_f&fid=37118&url=http%3A%2F%2Fwww.springerprotocols.com%2FAbstract%2Fdoi%2F10.1007%2F978-1-60761-580-4_1 In this chapter, we cover basic and fundamental principles and methods in statistics – from “What are Data and Statistics?” to “ANOVA and linear regression,” which are the basis of any statistical thinking and undertaking. Readers can easily find the selected topics in most introductory statistics textbooks, but we have tried to assemble and structure them in a succinct and reader-friendly manner in a stand-alone chapter. This text has long been used in real classroom settings for both undergraduate and graduate students who do or do not major in statistical sciences. We hope that from this chapter, readers would understand the key statistical concepts and terminologies, how to design a study (experimental or observational), how to analyze the data (e.g., desc... Springer protocols feed by Bioinformatics news http://www.medworm.com/rss/comments.php?id=4060025 Mon, 25 Jan 2010 00:00:00 +0100 4060025 http://www.medworm.com/index.php?rid=3114768&cid=s_37118_79_f&fid=37118&url=http%3A%2F%2Fwww.springerprotocols.com%2FAbstract%2Fdoi%2F10.1007%2F978-1-60761-444-9_9 Proteomics has advanced in leaps and bounds over the past couple of decades. However, the continuing dependency of mass spectrometry-based protein identification on the searching of spectra against protein sequence databases limits many proteomics experiments. If there is no sequenced genome for a given species, then cross species proteomics is required, attempting to identify proteins across the species boundary, typically using the sequenced genome of a closely related species. Unlike sequence searching for homologues, the proteomics equivalent is confounded by small differences in amino acid sequences, leading to large differences in peptide masses; this renders mass matching of peptides and their product ions difficult. Therefore, the phylogenetic distance between the two species and t... Springer protocols feed by Bioinformatics news http://www.medworm.com/rss/comments.php?id=3114768 Thu, 12 Nov 2009 00:00:00 +0100 3114768 http://www.medworm.com/index.php?rid=3114767&cid=s_37118_79_f&fid=37118&url=http%3A%2F%2Fwww.springerprotocols.com%2FAbstract%2Fdoi%2F10.1007%2F978-1-60761-444-9_8 The review describes methods of de novo sequencing of peptides by mass spectrometry. De novo methods utilize computational approaches to deduce the sequence or partial sequence of peptides directly from the experimental MS/MS spectra. The concepts behind a number of de novo sequencing methods are discussed. The other approach to identify peptides by tandem mass spectrometry is to match the fragment ions with virtual peptide ions generated from a genomic or protein database. De novo methods are essential to identify proteins when the genomes are not known but they are also extremely useful even when the genomes are known since they are not affected by errors in a search database. Another advantage of de novo methods is that the partial sequence can be used to search for posttranslation modi... Springer protocols feed by Bioinformatics news http://www.medworm.com/rss/comments.php?id=3114767 Thu, 12 Nov 2009 00:00:00 +0100 3114767 http://www.medworm.com/index.php?rid=3114766&cid=s_37118_79_f&fid=37118&url=http%3A%2F%2Fwww.springerprotocols.com%2FAbstract%2Fdoi%2F10.1007%2F978-1-60761-444-9_7 This article provides a concise roadmap for the proteomics researchers to start using spectral library searching in their data analysis workflow. (Source: Springer protocols feed by Bioinformatics) Springer protocols feed by Bioinformatics news http://www.medworm.com/rss/comments.php?id=3114766 Thu, 12 Nov 2009 00:00:00 +0100 3114766 http://www.medworm.com/index.php?rid=3114765&cid=s_37118_79_f&fid=37118&url=http%3A%2F%2Fwww.springerprotocols.com%2FAbstract%2Fdoi%2F10.1007%2F978-1-60761-444-9_6 Tandem mass spectrometry is a widely used tool in proteomics. This section will address the properties that describe how protonated peptides fragment when activated by collisions in a mass spectrometer and how that information can be used to identify proteins. A review of the mobile proton model is presented, along with a summary of commonly observed peptide cleavage enhancements, including the proline effect. The methods used to elucidate peptide dissociation chemistry by using both small groups of model peptides and large datasets are also discussed. Finally, the role of peak intensity in commercially available and developmental peptide identification algorithms is examined. (Source: Springer protocols feed by Bioinformatics) Springer protocols feed by Bioinformatics news http://www.medworm.com/rss/comments.php?id=3114765 Thu, 12 Nov 2009 00:00:00 +0100 3114765 http://www.medworm.com/index.php?rid=3114764&cid=s_37118_79_f&fid=37118&url=http%3A%2F%2Fwww.springerprotocols.com%2FAbstract%2Fdoi%2F10.1007%2F978-1-60761-444-9_5 Accurate and precise methods for estimating incorrect peptide and protein identifications are crucial for effective large-scale proteome analyses by tandem mass spectrometry. The target-decoy search strategy has emerged as a simple, effective tool for generating such estimations. This strategy is based on the premise that obvious, necessarily incorrect “decoy” sequences added to the search space will correspond with incorrect search results that might otherwise be deemed to be correct. With this knowledge, it is possible not only to estimate how many incorrect results are in a final data set but also to use decoy hits to guide the design of filtering criteria that sensitively partition a data set into correct and incorrect identifications. (Source: Springer protocols feed by Bi... Springer protocols feed by Bioinformatics news http://www.medworm.com/rss/comments.php?id=3114764 Thu, 12 Nov 2009 00:00:00 +0100 3114764 http://www.medworm.com/index.php?rid=3114763&cid=s_37118_79_f&fid=37118&url=http%3A%2F%2Fwww.springerprotocols.com%2FAbstract%2Fdoi%2F10.1007%2F978-1-60761-444-9_4 A variety of methods are described in the literature to assign peptide sequences to observed tandem MS data. Typically, the identified peptides are associated only with an arbitrary score that reflects the quality of the peptide-spectrum match but not with a statistically meaningful significance measure. In this chapter, we discuss why statistical significance measures can simplify and unify the interpretation of MS-based proteomic experiments. In addition, we also present available software solutions that convert scores into sound statistical measures. (Source: Springer protocols feed by Bioinformatics) Springer protocols feed by Bioinformatics news http://www.medworm.com/rss/comments.php?id=3114763 Thu, 12 Nov 2009 00:00:00 +0100 3114763 http://www.medworm.com/index.php?rid=3114762&cid=s_37118_79_f&fid=37118&url=http%3A%2F%2Fwww.springerprotocols.com%2FAbstract%2Fdoi%2F10.1007%2F978-1-60761-444-9_3 The peptide identification problem lies at the heart of modern proteomic methodology, from which the presence of a particular protein or proteins in a sample may be inferred. The challenge is to find the most likely amino acid sequence, which corresponds to each tandem mass spectrum that has been collected, and produce some kind of score and associated statistical measure that the putative identification is correct. This approach assumes that the peptide (and parent protein) sequence in question is known and is present in the database which is to be searched, as opposed to de novo methods, which seek to identify the peptide ab initio. This chapter will provide an overview of the methods that common, popular software tools employ to search protein sequence databases to provide the non-exper... Springer protocols feed by Bioinformatics news http://www.medworm.com/rss/comments.php?id=3114762 Thu, 12 Nov 2009 00:00:00 +0100 3114762 http://www.medworm.com/index.php?rid=3114761&cid=s_37118_79_f&fid=37118&url=http%3A%2F%2Fwww.springerprotocols.com%2FAbstract%2Fdoi%2F10.1007%2F978-1-60761-444-9_2 Protein identification by mass spectrometry (MS) is an important technique in proteomics. By searching an MS spectrum against a given protein database, the most matched proteins are sorted using a scoring function and the top one is often considered the correctly identified protein. Peptide mass fingerprinting (PMF) is one of the major methods for protein identification using MS technology. It is faster and cheaper than the other popular technique - Tandem Mass Spectrometry. Key bioinformatics issues in PMF analysis include designing a scoring function to quantitatively measure the degree of consistency between a PMF spectrum and a protein sequence and assessing the confidence of identified proteins. In this chapter, we will introduce several scoring functions that were developed by others... Springer protocols feed by Bioinformatics news http://www.medworm.com/rss/comments.php?id=3114761 Thu, 12 Nov 2009 00:00:00 +0100 3114761 http://www.medworm.com/index.php?rid=3114760&cid=s_37118_79_f&fid=37118&url=http%3A%2F%2Fwww.springerprotocols.com%2FAbstract%2Fdoi%2F10.1007%2F978-1-60761-444-9_25 Proteomics attempts to characterise the gene products expressed in a cell or tissue via a range of biophysical techniques including crystallography and NMR and, more relevantly to this volume, chromatography and mass spectrometry. It is becoming increasingly clear that the native states of segments of many of the cellular proteins are not stable, folded structures, and much of the proteome is in an unfolded, disordered state. These proteins and their disordered segments have functionally interesting properties and provide novel challenges for the biophysical techniques that are used to study them. This chapter focuses on computational approaches to predicting such regions and analyzing the functions linked to them, and has implications for protein scientists who wish to study such properti... Springer protocols feed by Bioinformatics news http://www.medworm.com/rss/comments.php?id=3114760 Thu, 12 Nov 2009 00:00:00 +0100 3114760 http://www.medworm.com/index.php?rid=3114759&cid=s_37118_79_f&fid=37118&url=http%3A%2F%2Fwww.springerprotocols.com%2FAbstract%2Fdoi%2F10.1007%2F978-1-60761-444-9_24 In Proteomics, fast enhancements with regard to technology are responsible for the creation of huge data sets. Consequently, in 2006 the European Commission funded a Coordination Action named ProDaC (Proteomics Data Collection) within the 6th EU Framework Programme to foster a community-wide data collection and data sharing. The aims of ProDaC were the development of documentation and storage standards, setup of a standardized data submission pipeline and collection of data. (Source: Springer protocols feed by Bioinformatics) Springer protocols feed by Bioinformatics news http://www.medworm.com/rss/comments.php?id=3114759 Thu, 12 Nov 2009 00:00:00 +0100 3114759 http://www.medworm.com/index.php?rid=3114758&cid=s_37118_79_f&fid=37118&url=http%3A%2F%2Fwww.springerprotocols.com%2FAbstract%2Fdoi%2F10.1007%2F978-1-60761-444-9_23 Data management and sharing in omics science is highly challenging due to the constant evolution of experimental techniques, the range of instrument types and software used for analysis, and the high volumes of data produced. The Functional Genomics Experiment (FuGE) Model was created to provide a model for capturing descriptions of sample processing, experimental protocols and multidimensional data for any kind of omics experiment. FuGE has two modes of action: (a) as a storage architecture for experimental workflows and (b) as a framework for building new technology-specific data standards. (Source: Springer protocols feed by Bioinformatics) Springer protocols feed by Bioinformatics news http://www.medworm.com/rss/comments.php?id=3114758 Thu, 12 Nov 2009 00:00:00 +0100 3114758 http://www.medworm.com/index.php?rid=3114757&cid=s_37118_79_f&fid=37118&url=http%3A%2F%2Fwww.springerprotocols.com%2FAbstract%2Fdoi%2F10.1007%2F978-1-60761-444-9_22 Mass spectrometry is an important technique for analyzing proteins and other biomolecular compounds in biological samples. Each of the vendors of these mass spectrometers uses a different proprietary binary output file format, which has hindered data sharing and the development of open source software for downstream analysis. The solution has been to develop, with the full participation of academic researchers as well as software and hardware vendors, an open XML-based format for encoding mass spectrometer output files, and then to write software to use this format for archiving, sharing, and processing. This chapter presents the various components and information available for this format, mzML. In addition to the XML schema that defines the file structure, a controlled vocabulary provide... Springer protocols feed by Bioinformatics news http://www.medworm.com/rss/comments.php?id=3114757 Thu, 12 Nov 2009 00:00:00 +0100 3114757 http://www.medworm.com/index.php?rid=3114756&cid=s_37118_79_f&fid=37118&url=http%3A%2F%2Fwww.springerprotocols.com%2FAbstract%2Fdoi%2F10.1007%2F978-1-60761-444-9_21 Molecular interactions are crucial components of the cellular process. In order to understand this complex machinery, one needs to gather published data from various sources. Many projects have initiated the collection of interaction data for this purpose since 2002. However, the lack of standardisation previously made the task of aggregating datasets difficult. This issue has been resolved by the creation of Molecular Interaction standard in 2004 by members of the Proteomics Standards Initiative (PSI), a work group of the Human Proteome Organization (HUPO). Furthermore, major database providers have come together with the goal to exchange data in order to optimise laborious curation tasks. Finally, tools and frameworks have been created based on PSI-MI standards to facilitate the visualis... Springer protocols feed by Bioinformatics news http://www.medworm.com/rss/comments.php?id=3114756 Thu, 12 Nov 2009 00:00:00 +0100 3114756 http://www.medworm.com/index.php?rid=3114755&cid=s_37118_79_f&fid=37118&url=http%3A%2F%2Fwww.springerprotocols.com%2FAbstract%2Fdoi%2F10.1007%2F978-1-60761-444-9_20 The PRIDE Proteomics Identifications Database provides users with the ability to explore and compare mass spectrometry-based proteomics experiments that reveal details of the protein expression found in a broad range of taxonomic groups, tissues and disease states. A PRIDE experiment typically includes identifications of proteins, peptides and protein modifications. Many of the submitted experiments also include processed peak lists representing the mass spectra that provide the evidence for these identifications. (Source: Springer protocols feed by Bioinformatics) Springer protocols feed by Bioinformatics news http://www.medworm.com/rss/comments.php?id=3114755 Thu, 12 Nov 2009 00:00:00 +0100 3114755 http://www.medworm.com/index.php?rid=3114754&cid=s_37118_79_f&fid=37118&url=http%3A%2F%2Fwww.springerprotocols.com%2FAbstract%2Fdoi%2F10.1007%2F978-1-60761-444-9_1 This book is part of the Methods in Molecular Biology series, and provides a general overview of computational approaches used in proteome research. In this chapter, we give an overview of the scope of the book in terms of current proteomics experimental techniques and the reasons why computational approaches are needed. We then give a summary of each chapter, which together provide a picture of the state of the art in proteome bioinformatics research. (Source: Springer protocols feed by Bioinformatics) Springer protocols feed by Bioinformatics news http://www.medworm.com/rss/comments.php?id=3114754 Thu, 12 Nov 2009 00:00:00 +0100 3114754 http://www.medworm.com/index.php?rid=3114753&cid=s_37118_79_f&fid=37118&url=http%3A%2F%2Fwww.springerprotocols.com%2FAbstract%2Fdoi%2F10.1007%2F978-1-60761-444-9_19 PeptideAtlas is a multi-species compendium of peptides observed with tandem mass spectrometry methods. Raw mass spectrometer output files are collected from the community and reprocessed through a uniform analysis and validation pipeline that continues to advance. The results are loaded into a database and the information derived from the raw data is returned to the community via several web-based data exploration tools. The PeptideAtlas resource is useful for experiment planning, improving genome annotation, and other data mining projects. PeptideAtlas has become especially useful for planning targeted proteomics experiments. (Source: Springer protocols feed by Bioinformatics) Springer protocols feed by Bioinformatics news http://www.medworm.com/rss/comments.php?id=3114753 Thu, 12 Nov 2009 00:00:00 +0100 3114753 http://www.medworm.com/index.php?rid=3114752&cid=s_37118_79_f&fid=37118&url=http%3A%2F%2Fwww.springerprotocols.com%2FAbstract%2Fdoi%2F10.1007%2F978-1-60761-444-9_18 Protein quantification represents an important extension to identification proteomics, enabling the comparison of protein expression across different samples or treatments. Comparative protein quantification by mass spectrometry typically employs stable isotope incorporation, but recently, comparative quantification of label-free LCn-MS proteomics data has emerged as an alternative approach. In this chapter, we provide an overview of the different approaches for extracting quantitative data from label-free LCn-MS experiments. The computational procedure for recovering the quantitative information is outlined. Examples of statistical tests used to evaluate the relevance of results are also provided. (Source: Springer protocols feed by Bioinformatics) Springer protocols feed by Bioinformatics news http://www.medworm.com/rss/comments.php?id=3114752 Thu, 12 Nov 2009 00:00:00 +0100 3114752 http://www.medworm.com/index.php?rid=3114751&cid=s_37118_79_f&fid=37118&url=http%3A%2F%2Fwww.springerprotocols.com%2FAbstract%2Fdoi%2F10.1007%2F978-1-60761-444-9_17 Isotope labeling combined with LC-MS/MS provides a robust platform for quantitative proteomics. Protein quantitation based on mass spectral data falls into two categories: one determined by MS/MS scans, e.g., iTRAQ-labeling quantitation, and the other by MS scans, e.g., quantitation using SILAC, ICAT, or 18O labeling. In large-scale LC-MS proteomic experiments, tens of thousands of MS and MS/MS spectra are generated and need to be analyzed. Data noise further complicates the data analysis. In this chapter, we present two automated tools, called Multi-Q and MaXIC-Q, for MS/MS- and MS-based quantitation analysis. They are designed as generic platforms that can accommodate search results from SEQUEST and Mascot, as well as mzXML files converted from raw files produced by various mass spectrom... Springer protocols feed by Bioinformatics news http://www.medworm.com/rss/comments.php?id=3114751 Thu, 12 Nov 2009 00:00:00 +0100 3114751 http://www.medworm.com/index.php?rid=3114750&cid=s_37118_79_f&fid=37118&url=http%3A%2F%2Fwww.springerprotocols.com%2FAbstract%2Fdoi%2F10.1007%2F978-1-60761-444-9_16 Despite recent progress in “shotgun” peptide separation by integrated liquid chromatography and mass spectrometry (LC/MS), proteome coverage and reproducibility are still limited with this approach and obtaining enough replicate runs for biomarker discovery is a challenge. For these reasons, recent research demonstrates that there is a continuing need for protein separation by two-dimensional gel electrophoresis (2-DE). However, with traditional 2-DE informatics, the digitized images are reduced to symbolic data through spot detection and quantification before proteins are compared for differential expression by spot matching. Recently, a more robust and automated paradigm has emerged where gels are directly aligned in the image domain before spots are detected across the whole... Springer protocols feed by Bioinformatics news http://www.medworm.com/rss/comments.php?id=3114750 Thu, 12 Nov 2009 00:00:00 +0100 3114750 http://www.medworm.com/index.php?rid=3114749&cid=s_37118_79_f&fid=37118&url=http%3A%2F%2Fwww.springerprotocols.com%2FAbstract%2Fdoi%2F10.1007%2F978-1-60761-444-9_15 Mass spectrometry has quickly become an essential tool in molecular biology laboratories. Here, we describe the Trans-Proteomic Pipeline, a collection of software tools, to facilitate the analysis, exchange, and comparison of MS data. The pipeline is instrument-independent and supports most commonly used proteomics workflows, including quantitative applications such as ICAT, iTRAQ, and SILAC. Importantly, the pipeline uses open, standard data formats and calculates accurate estimates of sensitivity and error rates, thus allowing for meaningful data exchange. In this chapter, we will introduce the various components of the pipeline in the context of three typical proteomic use-case scenarios. (Source: Springer protocols feed by Bioinformatics) Springer protocols feed by Bioinformatics news http://www.medworm.com/rss/comments.php?id=3114749 Thu, 12 Nov 2009 00:00:00 +0100 3114749 http://www.medworm.com/index.php?rid=3114748&cid=s_37118_79_f&fid=37118&url=http%3A%2F%2Fwww.springerprotocols.com%2FAbstract%2Fdoi%2F10.1007%2F978-1-60761-444-9_14 We describe the overall concepts behind the software and illustrate its use with several examples. (Source: Springer protocols feed by Bioinformatics) Springer protocols feed by Bioinformatics news http://www.medworm.com/rss/comments.php?id=3114748 Thu, 12 Nov 2009 00:00:00 +0100 3114748 http://www.medworm.com/index.php?rid=3114747&cid=s_37118_79_f&fid=37118&url=http%3A%2F%2Fwww.springerprotocols.com%2FAbstract%2Fdoi%2F10.1007%2F978-1-60761-444-9_13 Multiple reaction monitoring (MRM) of peptides is a popular proteomics technique that employs tandem mass spectrometry to quantify selected proteins of interest, such as those previously identified in differential protein identification studies. Using this technique, the specificity of precursor to product transitions is exploited to determine the absolute quantity of multiple proteins in a single sample. Selection of suitable transitions is critical for the success of MRM experiments, but accurate theoretical prediction of fragmentation patterns and peptide signal intensity is currently not possible. A recently proposed solution to this problem is to combine knowledge of the preferred properties of transitions for MRM, taken from expert practitioners, with MS/MS evidence extracted from a ... Springer protocols feed by Bioinformatics news http://www.medworm.com/rss/comments.php?id=3114747 Thu, 12 Nov 2009 00:00:00 +0100 3114747 http://www.medworm.com/index.php?rid=3114746&cid=s_37118_79_f&fid=37118&url=http%3A%2F%2Fwww.springerprotocols.com%2FAbstract%2Fdoi%2F10.1007%2F978-1-60761-444-9_12 High-throughput, MS-based proteomics studies are generating very large volumes of biologically relevant data. Given the central role of proteomics in emerging fields such as system/synthetic biology and biomarker discovery, the amount of proteomic data is expected to grow at unprecedented rates over the next decades. At the moment, there is pressing need for high-performance computational solutions to accelerate the analysis and interpretation of this data. (Source: Springer protocols feed by Bioinformatics) Springer protocols feed by Bioinformatics news http://www.medworm.com/rss/comments.php?id=3114746 Thu, 12 Nov 2009 00:00:00 +0100 3114746 http://www.medworm.com/index.php?rid=3114745&cid=s_37118_79_f&fid=37118&url=http%3A%2F%2Fwww.springerprotocols.com%2FAbstract%2Fdoi%2F10.1007%2F978-1-60761-444-9_11 Computational proteomics applications are often imagined as a pipeline, where information is processed in each stage before it flows to the next one. Independent of the type of application, the first stage invariably consists of obtaining the raw mass spectrometric data from the spectrometer and preparing it for use in the later stages by enhancing the signal of interest while suppressing spurious components. Numerous approaches for preprocessing MS data have been described in the literature. In this chapter, we will describe both, standard techniques originating from classical signal and image processing, and novel computational approaches specifically tailored to the analysis of MS data sets. We will focus on low level signal processing tasks such as baseline reduction, denoising, and fe... Springer protocols feed by Bioinformatics news http://www.medworm.com/rss/comments.php?id=3114745 Thu, 12 Nov 2009 00:00:00 +0100 3114745 http://www.medworm.com/index.php?rid=3114744&cid=s_37118_79_f&fid=37118&url=http%3A%2F%2Fwww.springerprotocols.com%2FAbstract%2Fdoi%2F10.1007%2F978-1-60761-444-9_10 The utility of a genome sequence in biological research depends entirely on the comprehensive description of all of its functional elements. Analysis of genome sequences is still predominantly gene-centric (i.e., identifying gene models/open reading frames). In this article, we describe a proteomics-based method for identifying open reading frames that are missed by computational algorithms. Mass spectrometry-based identification of peptides and proteins from biological samples provide evidence for the expression of the genome sequence at the protein level. This proteogenomic annotation method combines computationally predicted ORFs and the genome sequence with proteomics to identify novel gene models. We also describe our proteogenomic mapping pipeline - a set of computational tools that ... Springer protocols feed by Bioinformatics news http://www.medworm.com/rss/comments.php?id=3114744 Thu, 12 Nov 2009 00:00:00 +0100 3114744 http://www.medworm.com/index.php?rid=2819371&cid=s_37118_79_f&fid=37118&url=http%3A%2F%2Fwww.springerprotocols.com%2FAbstract%2Fdoi%2F10.1007%2F978-1-59745-524-4_9 Ever-increasing size of the biomedical literature makes more precise information retrieval and tapping into implicit knowledge in scientific literature a necessity. In this chapter, first, three new variants of the expectation–maximization (EM) method for semisupervised document classification (Machine Learning 39:103–134, 2000) are introduced to refine biomedical literature meta-searches. The retrieval performance of a multi-mixture per class EM variant with Agglomerative Information Bottleneck clustering (Slonim and Tishby (1999) Agglomerative information bottleneck. In Proceedings of NIPS-12) using Davies–Bouldin cluster validity index (IEEE Transactions on Pattern Analysis and Machine Intelligence 1:224–227, 1979), rivaled the state-of-the-art transductive suppo... Springer protocols feed by Bioinformatics news http://www.medworm.com/rss/comments.php?id=2819371 Mon, 03 Aug 2009 23:00:00 +0100 2819371 http://www.medworm.com/index.php?rid=2819370&cid=s_37118_79_f&fid=37118&url=http%3A%2F%2Fwww.springerprotocols.com%2FAbstract%2Fdoi%2F10.1007%2F978-1-59745-524-4_8 In this chapter, we will focus on the role of bioinformatics to analyze a protein after its protein structure has been determined. First, we present how to validate protein structures for quality assurance. Then, we discuss how to analyze protein–protein interfaces and how to predict the biomolecule which is the biological oligomeric state of the protein. Finally, we discuss how to search for homologs based on the 3-D structure which is an essential step for understanding protein function. (Source: Springer protocols feed by Bioinformatics) Springer protocols feed by Bioinformatics news http://www.medworm.com/rss/comments.php?id=2819370 Mon, 03 Aug 2009 23:00:00 +0100 2819370 http://www.medworm.com/index.php?rid=2819369&cid=s_37118_79_f&fid=37118&url=http%3A%2F%2Fwww.springerprotocols.com%2FAbstract%2Fdoi%2F10.1007%2F978-1-59745-524-4_7 The observation that similar protein sequences fold into similar three-dimensional structures provides a basis for the methods which predict structural features of a novel protein based on the similarity between its sequence and sequences of known protein structures. Similarity over entire sequence or large sequence fragment(s) enables prediction and modeling of entire structural domains while statistics derived from distributions of local features of known protein structures make it possible to predict such features in proteins with unknown structures. The accuracy of models of protein structures is sufficient for many practical purposes such as analysis of point mutation effects, enzymatic reactions, interaction interfaces of protein complexes, and active sites. Protein models are also u... Springer protocols feed by Bioinformatics news http://www.medworm.com/rss/comments.php?id=2819369 Mon, 03 Aug 2009 23:00:00 +0100 2819369 http://www.medworm.com/index.php?rid=2819368&cid=s_37118_79_f&fid=37118&url=http%3A%2F%2Fwww.springerprotocols.com%2FAbstract%2Fdoi%2F10.1007%2F978-1-59745-524-4_6 Interest in simulation of large-scale metabolic networks, species development, and genesis of various diseases requires new simulation techniques to accommodate the high complexity of realistic biological networks. Information geometry and topological formalisms are proposed to analyze information processes. We analyze the complexity of large-scale biological networks as well as transition of the system functionality due to modification in the system architecture, system environment, and system components. (Source: Springer protocols feed by Bioinformatics) Springer protocols feed by Bioinformatics news http://www.medworm.com/rss/comments.php?id=2819368 Mon, 03 Aug 2009 23:00:00 +0100 2819368 http://www.medworm.com/index.php?rid=2819367&cid=s_37118_79_f&fid=37118&url=http%3A%2F%2Fwww.springerprotocols.com%2FAbstract%2Fdoi%2F10.1007%2F978-1-59745-524-4_5 Discovery of DNA sequence variants responsible for human phenotypic variation is key to advances in molecular diagnostics and medicines. Historically, variants that alter the protein-coding sequence of genes have been targeted when attempting to identify a trait’s etiology; this is done because the rules governing these regions are generally well-understood and candidate variants can be easily selected. However, the effects of variants on gene regulation are increasingly regarded as being as important as protein-coding variation in uncovering the nature of phenotypic variation. I discuss resources and methodology that have recently been developed to computationally prioritize variants that may alter gene expression. (Source: Springer protocols feed by Bioinformatics) Springer protocols feed by Bioinformatics news http://www.medworm.com/rss/comments.php?id=2819367 Mon, 03 Aug 2009 23:00:00 +0100 2819367 http://www.medworm.com/index.php?rid=2819366&cid=s_37118_79_f&fid=37118&url=http%3A%2F%2Fwww.springerprotocols.com%2FAbstract%2Fdoi%2F10.1007%2F978-1-59745-524-4_4 A famous joke story that exhibits the traditionally awkward alliance between theory and experiment and showing the differences between experimental biologists and theoretical modelers is when a University sends a biologist, a mathematician, a physicist, and a computer scientist to a walking trip in an attempt to stimulate interdisciplinary research. During a break, they watch a cow in a field nearby and the leader of the group asks, “I wonder how one could decide on the size of a cow?” Since a cow is a biological object, the biologist responded first: “I have seen many cows in this area and know it is a big cow.” The mathematician argued, “The true volume is determined by integrating the mathematical function that describes the outer surface of the cow’s... Springer protocols feed by Bioinformatics news http://www.medworm.com/rss/comments.php?id=2819366 Mon, 03 Aug 2009 23:00:00 +0100 2819366 http://www.medworm.com/index.php?rid=2819365&cid=s_37118_79_f&fid=37118&url=http%3A%2F%2Fwww.springerprotocols.com%2FAbstract%2Fdoi%2F10.1007%2F978-1-59745-524-4_3 This paper presents current progress in the development of semantic data integration environment which is a part of the Biomedical Informatics Research Network (BIRN; http://www.nbirn.net ) project. BIRN is sponsored by the National Center for Research Resources (NCRR), a component of the National Institutes of Health (NIH). A goal is the development of a cyberinfrastructure for biomedical research that supports advance data acquisition, data storage, data management, data integration, data mining, data visualization, and other computing and information processing services over the Internet. Each participating institution maintains storage of their experimental or computationally derived data. Mediator-based data integration system performs semantic integration ove... Springer protocols feed by Bioinformatics news http://www.medworm.com/rss/comments.php?id=2819365 Mon, 03 Aug 2009 23:00:00 +0100 2819365 http://www.medworm.com/index.php?rid=2819364&cid=s_37118_79_f&fid=37118&url=http%3A%2F%2Fwww.springerprotocols.com%2FAbstract%2Fdoi%2F10.1007%2F978-1-59745-524-4_2 To promote scientific discovery and education, the federated Biomedical Informatics Research Network (BIRN) Data Repository (BDR) supports data storage, sharing, querying, and downloading for the biomedical community, enabling the integration of multiple data resources from a single entry point. The BDR encourages data sharing both for investigators requesting assistance with databasing and informatics infrastructure, and for those wishing to extend the reach of existing data resources to be registered with the BDR. Both approaches rely heavily on data integration and knowledge management techniques, ensuring capabilities for intelligent exploration of diverse data resources that make up the BDR’s shared environment. Although the development of the BDR has been driven by BIRN testbed... Springer protocols feed by Bioinformatics news http://www.medworm.com/rss/comments.php?id=2819364 Mon, 03 Aug 2009 23:00:00 +0100 2819364 http://www.medworm.com/index.php?rid=2819363&cid=s_37118_79_f&fid=37118&url=http%3A%2F%2Fwww.springerprotocols.com%2FAbstract%2Fdoi%2F10.1007%2F978-1-59745-524-4_1 Organizing and annotating biomedical data in structured ways has gained much interest and focus in the last 30 years. Driven by decreases in digital storage costs and advances in genetics sequencing, imaging, electronic data collection, and microarray technologies, data is being collected at an alarming rate. The specialization of fields in biology and medicine demonstrates the need for somewhat different structures for storage and retrieval of data. For biologists, the need for structured information and integration across a number of domains drives development. For clinical researchers and hospitals, the need for a structured medical record accessible to, ideally, any medical practitioner who might require it during the course of research or patient treatment, patient confidentiality, an... Springer protocols feed by Bioinformatics news http://www.medworm.com/rss/comments.php?id=2819363 Mon, 03 Aug 2009 23:00:00 +0100 2819363 http://www.medworm.com/index.php?rid=2819362&cid=s_37118_79_f&fid=37118&url=http%3A%2F%2Fwww.springerprotocols.com%2FAbstract%2Fdoi%2F10.1007%2F978-1-59745-524-4_12 Single Sign-On (SSO) is a practical requirement for software applications, which rely on distributed, networked services requiring authentication. SSO is as much a convenient feature for users as it is a security concern for application designers. The security requirement becomes critical in institutions that adhere to HIPPA regulations. In this chapter, we discuss SSO as it applies to a grid portal using remote computational resources and grid storage, which contain Personal Health Information (PHI). We cover the implementation of Public Key Infrastructure(PKI) to meet HIPPA security requirements such as authentication, confidentiality, nonrepudiation, and dataintegrity. Furthermore, we discuss the different technologies in PKI that solves these security concerns with respect to protectin... Springer protocols feed by Bioinformatics news http://www.medworm.com/rss/comments.php?id=2819362 Mon, 03 Aug 2009 23:00:00 +0100 2819362 http://www.medworm.com/index.php?rid=2819361&cid=s_37118_79_f&fid=37118&url=http%3A%2F%2Fwww.springerprotocols.com%2FAbstract%2Fdoi%2F10.1007%2F978-1-59745-524-4_11 The field of high-performance computing (HPC) has provided a wide array of strategies for supplying additional computing power to the goal of reducing the total “clock time” required to complete various computational processes. These strategies range from the development of higher-performance hardware to the assembly of large networks of commodity computers, with each strategy designed to address a particular aspect and/or manifestation of a given computational problem. GWE (Grid Wizard Enterprise) in that regard, is an HPC distributed enterprise system, aimed at providing a solution to the particular problem of running inter-independent computational processes faster by parallelizing their execution across a virtual grid of computational resources with a minimum of user interv... Springer protocols feed by Bioinformatics news http://www.medworm.com/rss/comments.php?id=2819361 Mon, 03 Aug 2009 23:00:00 +0100 2819361 http://www.medworm.com/index.php?rid=2819360&cid=s_37118_79_f&fid=37118&url=http%3A%2F%2Fwww.springerprotocols.com%2FAbstract%2Fdoi%2F10.1007%2F978-1-59745-524-4_10 Imagination is the critical point in developing of realistic intelligence (AI) systems. One way to approach imagination would be simulation of its properties and operations. We developed two models “Brain Network Hierarchy of Languages,” and “Semantical Holographic Calculus” and simulation system ScriptWriter that emulate the process of imagination through an automatic animation of English texts. The purpose of this paper is to demonstrate the model and present “ScriptWriter” system http://nvo.sdsc.edu/NVO/JCSG/get_SRB_mime_file2.cgi//home/tamara.sdsc/test/demo.zip?F=/home/tamara.sdsc/test/demo.zip&M=application/x-gtar for simulation of the imagination. (Source: Springer protocols feed by Bioinformatics) Springer protocols feed by Bioinformatics news http://www.medworm.com/rss/comments.php?id=2819360 Mon, 03 Aug 2009 23:00:00 +0100 2819360 http://www.medworm.com/index.php?rid=2364082&cid=s_37118_79_f&fid=37118&url=http%3A%2F%2Fwww.springerprotocols.com%2FAbstract%2Fdoi%2F10.1007%2F978-1-59745-251-9_17 The analysis of DNA sequence polymorphisms and SNPs (single nucleotide polymorphisms) can provide insights into the evolutionary forces acting on populations and species. Available population-genetic methods, and particularly those based on the coalescent theory, have become the primary framework to analyze such DNA polymorphism data. Here, I explain some essential analytical methods for interpreting DNA polymorphism data and also describe the basic functionalities of the DnaSP software. DnaSP is a multi-propose program that allows conducting exhaustive DNA polymorphism analysis using a graphical user-friendly interface. (Source: Springer protocols feed by Bioinformatics) Springer protocols feed by Bioinformatics news http://www.medworm.com/rss/comments.php?id=2364082 Thu, 01 Jan 2009 05:00:00 +0100 2364082 http://www.medworm.com/index.php?rid=2364081&cid=s_37118_79_f&fid=37118&url=http%3A%2F%2Fwww.springerprotocols.com%2FAbstract%2Fdoi%2F10.1007%2F978-1-59745-251-9_10 We present principles and practical procedures for using analyses of GC content and codon usage frequency to identify highly expressed or horizontally transferred genes and to study the relative contribution of different types of mutation to gene and genome composition. CodonExplorer’s combination of a user-friendly web interface and a comprehensive genomic database makes these diverse analyses fast and straightforward to perform. CodonExplorer is thus a powerful tool that facilitates and automates a wide range of compositional analyses. (Source: Springer protocols feed by Bioinformatics) Springer protocols feed by Bioinformatics news http://www.medworm.com/rss/comments.php?id=2364081 Thu, 01 Jan 2009 05:00:00 +0100 2364081 http://www.medworm.com/index.php?rid=2364080&cid=s_37118_79_f&fid=37118&url=http%3A%2F%2Fwww.springerprotocols.com%2FAbstract%2Fdoi%2F10.1007%2F978-1-59745-251-9_11 There is a standard genetic code that is used by most organisms, but exceptions exist in which particular codons are translated with a different meaning, i.e., as a different amino acid. The characterization of the genetic code of an organism is hence a key step for properly analyzing and translating its protein-coding genes. Such characterization is particularly important in the case of metazoan mitochondrial genomes for two reasons: first, many variant codes occur in them and second, mitochondrial data is frequently used for evolutionary studies. Variant codes are usually found by comparative sequence analyses. Given a protein alignment, if a particular codon for a given species occurs at positions in which a particular amino acid is frequently found in other species, then the most likel... Springer protocols feed by Bioinformatics news http://www.medworm.com/rss/comments.php?id=2364080 Thu, 01 Jan 2009 05:00:00 +0100 2364080 http://www.medworm.com/index.php?rid=2364079&cid=s_37118_79_f&fid=37118&url=http%3A%2F%2Fwww.springerprotocols.com%2FAbstract%2Fdoi%2F10.1007%2F978-1-59745-251-9_12 We present in this chapter a simple protocol mainly based on the combination of the program GeneID and other computational tools to annotate the location of a gene, which was previously annotated in D. melanogaster, in the recently assembled genome of D. yakuba. (Source: Springer protocols feed by Bioinformatics) Springer protocols feed by Bioinformatics news http://www.medworm.com/rss/comments.php?id=2364079 Thu, 01 Jan 2009 05:00:00 +0100 2364079 http://www.medworm.com/index.php?rid=2364078&cid=s_37118_79_f&fid=37118&url=http%3A%2F%2Fwww.springerprotocols.com%2FAbstract%2Fdoi%2F10.1007%2F978-1-59745-251-9_13 Reliable detection of cis-regulatory elements in promoter regions is a difficult and unsolved problem in computational biology. The intricacy of transcriptional regulation in higher eukaryotes, primarily in metazoans, could be a major driving force of organismal complexity. Eukaryotic genome annotations have improved greatly due to large-scale characterization of full-length cDNAs, transcriptional start sites (TSSs), and comparative genomics. Regulatory elements are identified in promoter regions using a variety of enumerative or alignment-based methods. Here we present a survey of recent computational methods for eukaryotic promoter analysis and describe the use of an alignment-based method implemented in the A-GLAM program. (Source: Springer protocols feed by Bioinformatics) Springer protocols feed by Bioinformatics news http://www.medworm.com/rss/comments.php?id=2364078 Thu, 01 Jan 2009 05:00:00 +0100 2364078 http://www.medworm.com/index.php?rid=2364077&cid=s_37118_79_f&fid=37118&url=http%3A%2F%2Fwww.springerprotocols.com%2FAbstract%2Fdoi%2F10.1007%2F978-1-59745-251-9_14 Genomic DNA is being sequenced and annotated at a rapid rate, with terabases of DNA currently deposited in GenBank and other repositories. Genome browsers provide an essential collection of resources to visualize and analyze chromosomal DNA. The University of California, Santa Cruz (UCSC) Genome Browser provides annotations from the level of single nucleotides to whole chromosomes for four dozen metazoan and other species. The Genome Browser may be used to address a wide range of problems in bioinformatics (e.g., sequence analysis), comparative genomics, and evolution. (Source: Springer protocols feed by Bioinformatics) Springer protocols feed by Bioinformatics news http://www.medworm.com/rss/comments.php?id=2364077 Thu, 01 Jan 2009 05:00:00 +0100 2364077 http://www.medworm.com/index.php?rid=2364076&cid=s_37118_79_f&fid=37118&url=http%3A%2F%2Fwww.springerprotocols.com%2FAbstract%2Fdoi%2F10.1007%2F978-1-59745-251-9_16 We present here a survey of two of the most readily available and widely used bioinformatics applications for the detection, characterization, and analysis of TE sequences in eukaryotic genomes: CENSOR and RepeatMasker. For each program, information on availability, input, output, and the algorithmic methods used is provided. Specific examples of the use of CENSOR and RepeatMasker are also described. CENSOR and RepeatMasker both rely on homology-based methods for the detection of TE sequences. There are several other classes of methods available for the analysis of repetitive DNA sequences including de novo methods that compare genomic sequences against themselves, class-specific methods that use structural characteristics of specific classes of elements to aid in their identification, and... Springer protocols feed by Bioinformatics news http://www.medworm.com/rss/comments.php?id=2364076 Thu, 01 Jan 2009 05:00:00 +0100 2364076 http://www.medworm.com/index.php?rid=2364075&cid=s_37118_79_f&fid=37118&url=http%3A%2F%2Fwww.springerprotocols.com%2FAbstract%2Fdoi%2F10.1007%2F978-1-59745-251-9_15 Molecular genetic markers represent one of the most powerful tools for the analysis of genomes and the association of heritable traits with underlying genetic variation. The development of high-throughput methods for the detection of single nucleotide polymorphisms (SNPs) and simple sequence repeats (SSRs) has led to a revolution in their use as molecular markers. The availability of large sequence data sets permits mining for these molecular markers, which may then be used for applications such as genetic trait mapping, diversity analysis and marker assisted selection in agriculture. Here we describe web-based automated methods for the discovery of SSRs using SSR taxonomy tree, the discovery of SNPs from sequence data using SNPServer and the identification of validated SNPs from within th... Springer protocols feed by Bioinformatics news http://www.medworm.com/rss/comments.php?id=2364075 Thu, 01 Jan 2009 05:00:00 +0100 2364075 http://www.medworm.com/index.php?rid=2364074&cid=s_37118_79_f&fid=37118&url=http%3A%2F%2Fwww.springerprotocols.com%2FAbstract%2Fdoi%2F10.1007%2F978-1-59745-251-9_1 Similarity searches are an essential component of most bioinformatic applications. They form the bases of structural motif identification, gene identification, and insights into functional associations. With the rapid increase in the available genetic data through a wide variety of databases, similarity searches are an essential tool for accessing these data in an informative and productive way. In this chapter, we provide an overview of similarity searching approaches, related databases, and parameter options to achieve the best results for a variety of applications. We then provide a worked example and some notes for consideration. (Source: Springer protocols feed by Bioinformatics) Springer protocols feed by Bioinformatics news http://www.medworm.com/rss/comments.php?id=2364074 Thu, 01 Jan 2009 05:00:00 +0100 2364074 http://www.medworm.com/index.php?rid=2364073&cid=s_37118_79_f&fid=37118&url=http%3A%2F%2Fwww.springerprotocols.com%2FAbstract%2Fdoi%2F10.1007%2F978-1-59745-251-9_2 OrthologID ( http://nypg.bio.nyu.edu/orthologid/ ) allows for the rapid and accurate identification of gene orthology within a character-based phylogenetic framework. The Web application has two functions – an orthologous group search and a query orthology classification. The former determines orthologous gene sets for complete genomes and identifies diagnostic characters that define each orthologous gene set; and the latter allows for the classification of unknown query sequences to orthology groups. The first module of the Web application, the gene family generator, uses an E-value based approach to sort genes into gene families. An alignment constructor then aligns members of gene families and the resulting gene family alignments are submitted to the tree b... Springer protocols feed by Bioinformatics news http://www.medworm.com/rss/comments.php?id=2364073 Thu, 01 Jan 2009 05:00:00 +0100 2364073 http://www.medworm.com/index.php?rid=2364072&cid=s_37118_79_f&fid=37118&url=http%3A%2F%2Fwww.springerprotocols.com%2FAbstract%2Fdoi%2F10.1007%2F978-1-59745-251-9_3 Multiple alignment of DNA sequences is an important step in various molecular biological analyses. As a large amount of sequence data is becoming available through genome and other large-scale sequencing projects, scalability, as well as accuracy, is currently required for a multiple sequence alignment (MSA) program. In this chapter, we outline the algorithms of an MSA program MAFFT and provide practical advice, focusing on several typical situations a biologist sometimes faces. For genome alignment, which is beyond the scope of MAFFT, we introduce two tools: TBA and MAUVE. (Source: Springer protocols feed by Bioinformatics) Springer protocols feed by Bioinformatics news http://www.medworm.com/rss/comments.php?id=2364072 Thu, 01 Jan 2009 05:00:00 +0100 2364072 http://www.medworm.com/index.php?rid=2364071&cid=s_37118_79_f&fid=37118&url=http%3A%2F%2Fwww.springerprotocols.com%2FAbstract%2Fdoi%2F10.1007%2F978-1-59745-251-9_4 Compositional heterogeneity is a poorly appreciated attribute of aligned nucleotide and amino acid sequences. It is a common property of molecular phylogenetic data, and it has been found to occur across sequences and/or across sites. Most molecular phylogenetic methods assume that the sequences have evolved under globally stationary, reversible, and homogeneous conditions, implying that the sequences should be compositionally homogeneous. The presence of the above-mentioned compositional heterogeneity implies that the sequences must have evolved under more general conditions than is commonly assumed. Consequently, there is a need for reliable methods to detect under what conditions alignments of nucleotides or amino acids may have evolved. In this chapter, we describe one such program. Se... Springer protocols feed by Bioinformatics news http://www.medworm.com/rss/comments.php?id=2364071 Thu, 01 Jan 2009 05:00:00 +0100 2364071 http://www.medworm.com/index.php?rid=2364070&cid=s_37118_79_f&fid=37118&url=http%3A%2F%2Fwww.springerprotocols.com%2FAbstract%2Fdoi%2F10.1007%2F978-1-59745-251-9_5 jModelTest is a bioinformatic tool for choosing among different models of nucleotide substitution. The program implements five different model selection strategies, including hierarchical and dynamical likelihood ratio tests (hLRT and dLRT), Akaike and Bayesian information criteria (AIC and BIC), and a performance-based decision theory method (DT). The output includes estimates of model selection uncertainty, parameter importances, and model-averaged parameter estimates, including model-averaged phylogenies. jModelTest is a Java program that runs under Mac OSX, Windows, and Unix systems with a Java Run Environment installed, and it can be freely downloaded from http://darwin.uvigo.es . (Source: Springer protocols feed by Bioinformatics) Springer protocols feed by Bioinformatics news http://www.medworm.com/rss/comments.php?id=2364070 Thu, 01 Jan 2009 05:00:00 +0100 2364070 http://www.medworm.com/index.php?rid=2364069&cid=s_37118_79_f&fid=37118&url=http%3A%2F%2Fwww.springerprotocols.com%2FAbstract%2Fdoi%2F10.1007%2F978-1-59745-251-9_6 Our understanding of the origins, the functions and/or the structures of biological sequences strongly depends on our ability to decipher the mechanisms of molecular evolution. These complex processes can be described through the comparison of homologous sequences in a phylogenetic framework. Moreover, phylogenetic inference provides sound statistical tools to exhibit the main features of molecular evolution from the analysis of actual sequences. This chapter focuses on phylogenetic tree estimation under the maximum likelihood (ML) principle. Phylogenies inferred under this probabilistic criterion are usually reliable and important biological hypotheses can be tested through the comparison of different models. Estimating ML phylogenies is computationally demanding, and careful examination ... Springer protocols feed by Bioinformatics news http://www.medworm.com/rss/comments.php?id=2364069 Thu, 01 Jan 2009 05:00:00 +0100 2364069 http://www.medworm.com/index.php?rid=2364068&cid=s_37118_79_f&fid=37118&url=http%3A%2F%2Fwww.springerprotocols.com%2FAbstract%2Fdoi%2F10.1007%2F978-1-59745-251-9_7 We describe the most widely used supertree methods implemented in the software program “clann” and provide a step by step tutorial for investigating phylogenetic information and reconstructing the best supertree. Clann is freely available for Windows, Mac and Unix/Linux operating systems under the GNU public licence at http://bioinf.nuim.ie/software/clann . (Source: Springer protocols feed by Bioinformatics) Springer protocols feed by Bioinformatics news http://www.medworm.com/rss/comments.php?id=2364068 Thu, 01 Jan 2009 05:00:00 +0100 2364068 http://www.medworm.com/index.php?rid=2364067&cid=s_37118_79_f&fid=37118&url=http%3A%2F%2Fwww.springerprotocols.com%2FAbstract%2Fdoi%2F10.1007%2F978-1-59745-251-9_8 Natural selection is a fundamental process affecting all evolving populations. In the simplest case, positive selection increases the frequency of alleles that confer a fitness advantage relative to the rest of the population, or increases its genetic diversity, and negative selection removes those alleles that are deleterious. Codon-based models of molecular evolution are able to infer signatures of selection from alignments of homologous sequences by estimating the relative rates of synonymous (dS) and non-synonymous substitutions (dN). Datamonkey ( http://www.datamonkey.org ) provides a user-friendly web interface to a wide collection of state-of-the-art statistical techniques for estimating dS and dN and identifying codons and lineages under selection, even in t... Springer protocols feed by Bioinformatics news http://www.medworm.com/rss/comments.php?id=2364067 Thu, 01 Jan 2009 05:00:00 +0100 2364067 http://www.medworm.com/index.php?rid=2364066&cid=s_37118_79_f&fid=37118&url=http%3A%2F%2Fwww.springerprotocols.com%2FAbstract%2Fdoi%2F10.1007%2F978-1-59745-251-9_9 Recombination between nucleotide sequences is a major process influencing the evolution of most species on Earth. While its evolutionary value is a matter of quite intense debate, so too is the influence of recombination on evolutionary analysis methods that assume nucleotide sequences replicate without recombining. The crux of the problem is that when nucleic acids recombine, the daughter or recombinant molecules no longer have a single evolutionary history. All analysis methods that derive increased power from correctly inferring evolutionary relationships between sequences will therefore be at least mildly sensitive to the effects of recombination. The importance of considering recombination in evolutionary studies is underlined by the bewildering array of currently available methods an... Springer protocols feed by Bioinformatics news http://www.medworm.com/rss/comments.php?id=2364066 Thu, 01 Jan 2009 05:00:00 +0100 2364066 http://www.medworm.com/index.php?rid=2364090&cid=s_37118_79_f&fid=37118&url=http%3A%2F%2Fwww.springerprotocols.com%2FAbstract%2Fdoi%2F10.1007%2F978-1-59745-243-4_17 Enzymes play central roles in metabolic pathways, and the prediction of metabolic pathways in newly sequenced genomes usually starts with the assignment of genes to enzymatic reactions. However, genes with similar catalytic activity are not necessarily similar in sequence, and therefore the traditional sequence similarity-based approach often fails to identify the relevant enzymes, thus hindering efforts to map the metabolome of an organism. (Source: Springer protocols feed by Bioinformatics) Springer protocols feed by Bioinformatics news http://www.medworm.com/rss/comments.php?id=2364090 Tue, 01 Jul 2008 04:00:00 +0100 2364090 http://www.medworm.com/index.php?rid=2364089&cid=s_37118_79_f&fid=37118&url=http%3A%2F%2Fwww.springerprotocols.com%2FAbstract%2Fdoi%2F10.1007%2F978-1-59745-243-4_18 Intricate networks of protein interactions rely on the ability of a protein to recognize its targets: other proteins, ligands, and sites on DNA and RNA. To recognize other molecules, it was suggested that a protein uses a small set of specificity-determining residues (SDRs). How can one find these residues in proteins and distinguish them from other functionally important amino acids? A number of bioinformatics methods to predict SDRs have been developed in recent years. These methods use genomic information and multiple sequence alignments to identify positions exhibiting a specific pattern of conservation and variability. The challenge is to delineate the evolutionary pattern of SDRs from that of the active site residues and the residues responsible for formation of the protein’s s... Springer protocols feed by Bioinformatics news http://www.medworm.com/rss/comments.php?id=2364089 Tue, 01 Jul 2008 04:00:00 +0100 2364089