Computational methods and tools are nowadays widely applied for rational Metabolic Engineering approaches. However, what is still missing are clear advices on the right order of the application of these tools. The ava...
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Computational methods and tools are nowadays widely applied for rational Metabolic Engineering approaches. However, what is still missing are clear advices on the right order of the application of these tools. The availability of genomic information for a large number of cellular systems especially requires the use of computers to store, analyze, and process knowledge of single enzymes, metabolic pathways, and cellular networks. The trend of integrating measured quantities for the metabolome, the transcriptome, and the proteome into mathematical models, combined with methods for the rational design of cellular networks, has led to the research field Systems Metabolic Engineering, a field that extends and amplifies the classical field of Metabolic Engineering. This chapter describes mathematical and computational approaches on the cellular and the process levels. In the Material section, modeling approaches and methods for model analysis are introduced, and the current state of the art is reviewed. In the Method section, we propose a protocol for efficiently combining various approaches for the optimal production of desired biotechnological products. less
The ability to extract, identify and annotate large amounts of biological data is a key feature of the “omics” era, and has led to an explosion in the amount of data available. One pivotal advance is the use of Next...
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The ability to extract, identify and annotate large amounts of biological data is a key feature of the “omics” era, and has led to an explosion in the amount of data available. One pivotal advance is the use of Next-Generation Sequencing (NGS) techniques such as RNA-sequencing (RNA-seq). RNA-seq uses data from millions of small mRNA transcripts or “reads” which are aligned to a reference genome. Comparative transcriptomics analyses using RNA-seq can provide the researcher with a comprehensive view of the cells’ response to a given environment or stimulus. Here, we describe the NGS techniques (based on Illumina technology) that are routinely used for comparative transcriptome analysis of fungal species. We describe the entire process from isolation of RNA to computational identification of differentially expressed genes. We provide instructions to allow the beginner to implement packages in R such as Bioconductor. The methods described are not limited to yeast, and can also be applied to other eukaryotic organisms. less
Parallel sequencing of a single cell's genome and transcriptome provides a powerful tool for dissecting genetic variation and its relationship with gene expression. Here we present a detailed protocol for G&T-...
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Parallel sequencing of a single cell's genome and transcriptome provides a powerful tool for dissecting genetic variation and its relationship with gene expression. Here we present a detailed protocol for G&T-seq, a method for separation and parallel sequencing of genomic DNA and full-length polyA(+) mRNA from single cells. We provide step-by-step instructions for the isolation and lysis of single cells; the physical separation of polyA(+) mRNA from genomic DNA using a modified oligo-dT bead capture and the respective whole-transcriptome and whole-genome amplifications; and library preparation and sequence analyses of these amplification products. The method allows the detection of thousands of transcripts in parallel with the genetic variants captured by the DNA-seq data from the same single cell. G&T-seq differs from other currently available methods for parallel DNA and RNA sequencing from single cells, as it involves physical separation of the DNA and RNA and does not require bespoke microfluidics platforms. The process can be implemented manually or through automation. When performed manually, paired genome and transcriptome sequencing libraries from eight single cells can be produced in ∼3 d by researchers experienced in molecular laboratory work. For users with experience in the programming and operation of liquid-handling robots, paired DNA and RNA libraries from 96 single cells can be produced in the same time frame. Sequence analysis and integration of single-cell G&T-seq DNA and RNA data requires a high level of bioinformatics expertise and familiarity with a wide range of informatics tools. less
The simulation of protein aggregation poses several computational challenges due to the disparate time and lengths scales that are involved. This chapter focuses on the use of atomistically detailed simulations to pro...
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The simulation of protein aggregation poses several computational challenges due to the disparate time and lengths scales that are involved. This chapter focuses on the use of atomistically detailed simulations to probe the initial steps of aggregation, with an emphasis on the Tau peptide as a model system, run under a replica exchange molecular dynamics protocol. less
Peptide microarrays are a powerful tool to identify linear epitopes of food allergens in a high-throughput manner. The main advantages of the microarray-based immunoassay are the possibility to assay thousands of targ...
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Peptide microarrays are a powerful tool to identify linear epitopes of food allergens in a high-throughput manner. The main advantages of the microarray-based immunoassay are the possibility to assay thousands of targets simultaneously, the requirement of a low volume of serum, the more robust statistical analysis, and the possibility to test simultaneously several immunoglobulin subclasses. Among them, the last one has a special interest in the field of food allergy, because the development of tolerance to food allergens has been associated with a decrease in IgE and an increase in IgG4 levels against linear epitopes. However, the main limitation to the clinical use of microarray is the automated analysis of the data. Recent studies mapping the linear epitopes of food allergens with peptide microarray immunoassays have identified peptide biomarkers that can be used for early diagnosis of food allergies and to predict their severity or the self-development of tolerance. Using this approach, we have worked on epitope mapping of the two most important food allergens in the Spanish population, cow’s milk and chicken eggs. The final aim of these studies is to define subsets of peptides that could be used as biomarkers to improve the diagnosis and prognosis of food allergies. This chapter describes the protocol to produce microarrays using a library of overlapping peptides corresponding to the primary sequences of food allergens and data acquisition and analysis of IgE- and IgG4-binding epitopes. less
Conservation of particular molecular sequence motifs throughout evolution is a strong indicator of their functional relevance as selective pressure likely prevented the accumulation of mutations. Known as “phylogenet...
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Conservation of particular molecular sequence motifs throughout evolution is a strong indicator of their functional relevance as selective pressure likely prevented the accumulation of mutations. Known as “phylogenetic footprinting”, this rationale has been exploited for the identification of novel functional motifs using sequence information from sequence alignments of diverse species, in particular transcription factor binding site motifs in aligned gene promoter sequences of orthologous genes. With the rapid advances of sequencing technologies, whole genome sequence information is accumulating not only across different species, but increasingly for variants of the same species exhibiting relatively little sequence variability, primarily present as single nucleotide polymorphisms (SNPs). Here, we lay out the basic strategy for the identification of functional cis-regulatory motifs in gene promoter regions based on SNP information. less
Connectomics—the study of how neurons wire together in the brain—is at the forefront of modern neuroscience research. However, many connectomics studies are limited by the time and precision needed to correctly segm...
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Connectomics—the study of how neurons wire together in the brain—is at the forefront of modern neuroscience research. However, many connectomics studies are limited by the time and precision needed to correctly segment large volumes of electron microscopy (EM) image data. We present here a semi-automated segmentation pipeline using freely available software that can significantly decrease segmentation time for extracting both nuclei and cell bodies from EM image volumes. less
An intelligent control system for an X ray streak camera in a diagnostic instrument manipulator (DIM) is proposed and implemented, which can control time delay, electric focusing, image gain adjustment, switch of swee...
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An intelligent control system for an X ray streak camera in a diagnostic instrument manipulator (DIM) is proposed and implemented, which can control time delay, electric focusing, image gain adjustment, switch of sweep voltage, acquiring environment parameters etc. The system consists of 16 A/D converters and 16 D/A converters, a 32-channel general purpose input/output (GPIO) and two sensors. An isolated DC/DC converter with multi-outputs and a single mode filer were adopted to reduce the interference generated by the common ground among the A/D, D/A and I/O. The software was designed using graphical programming language and can remotely access the corresponding instrument from a website. The entire intelligent control system can acquire the desirable data at a speed of 30 Mb/s and store 11 for later analysis. The intelligent. system was implemented on a streak camera in a DIM and it shows a temporal resolution of 11.25 ps, spatial distortion of less than 10% and dynamic range of 279:1. The intelligent control system has been successfully used in a streak camera to verify the synchronization of multi-channel laser on the Confinement Fusion (C) 2015 Elsevier B.V. All rights reserved.
With the advancement in proteomics separation techniques and improvements in mass analyzers, the data generated in a mass-spectrometry based proteomics experiment is rising exponentially. Such voluminous datasets nece...
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With the advancement in proteomics separation techniques and improvements in mass analyzers, the data generated in a mass-spectrometry based proteomics experiment is rising exponentially. Such voluminous datasets necessitate automated computational tools for high-throughput data analysis and appropriate statistical control. The data is searched using one or more of the several popular database search algorithms. The matches assigned by these tools can have false positives and statistical validation of these false matches is necessary before making any biological interpretations. Without such procedures, the biological inferences do not hold true and may be outright misleading. There is a considerable overlap between true and false positives. To control the false positives amongst a set of accepted matches, there is a need for some statistical estimate that can reflect the amount of false positives present in the data processed. False discovery rate (FDR) is the metric for global confidence assessment of a large-scale proteomics dataset. This chapter covers the basics of FDR, its application in proteomics, and methods to estimate FDR. less
The purpose of this thesis is to study the plagiarism detection method aiming at C++ programs. We proposed the corresponding preventive measures by summarizing the common types of plagiarism attack through observation...
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ISBN:
(纸本)9783642166952
The purpose of this thesis is to study the plagiarism detection method aiming at C++ programs. We proposed the corresponding preventive measures by summarizing the common types of plagiarism attack through observation and statistical analysis. Using text analysis, structure analysis, and variable analysis would prevent misjudgment. Finally, we implemented the CPD(C++ Plagiarism Detection) system and compare it with other existing systems, and the experimental result shows us that our system can detect more kinds of plagiarism attacks than other existing systems.
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