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1.  GACT: a Genome build and Allele definition Conversion Tool for SNP imputation and meta-analysis in genetic association studies 
BMC Genomics  2014;15:610.
Genome-wide association studies (GWAS) have successfully identified genes associated with complex human diseases. Although much of the heritability remains unexplained, combining single nucleotide polymorphism (SNP) genotypes from multiple studies for meta-analysis will increase the statistical power to identify new disease-associated variants. Meta-analysis requires same allele definition (nomenclature) and genome build among individual studies. Similarly, imputation, commonly-used prior to meta-analysis, requires the same consistency. However, the genotypes from various GWAS are generated using different genotyping platforms, arrays or SNP-calling approaches, resulting in use of different genome builds and allele definitions. Incorrect assumptions of identical allele definition among combined GWAS lead to a large portion of discarded genotypes or incorrect association findings. There is no published tool that predicts and converts among all major allele definitions.
In this study, we have developed a tool, GACT, which stands for Genome build and Allele definition Conversion Tool, that predicts and inter-converts between any of the common SNP allele definitions and between the major genome builds. In addition, we assessed several factors that may affect imputation quality, and our results indicated that inclusion of singletons in the reference had detrimental effects while ambiguous SNPs had no measurable effect. Unexpectedly, exclusion of genotypes with missing rate > 0.001 (40% of study SNPs) showed no significant decrease of imputation quality (even significantly higher when compared to the imputation with singletons in the reference), especially for rare SNPs.
GACT is a new, powerful, and user-friendly tool with both command-line and interactive online versions that can accurately predict, and convert between any of the common allele definitions and between genome builds for genome-wide meta-analysis and imputation of genotypes from SNP-arrays or deep-sequencing, particularly for data from the dbGaP and other public databases.
GACT software
PMCID: PMC4223508  PMID: 25038819
Allele definition (nomenclature); Genome build; Genome-wide association study (GWAS); Imputation; Meta-analysis
2.  RECLU: a pipeline to discover reproducible transcriptional start sites and their alternative regulation using capped analysis of gene expression (CAGE) 
BMC Genomics  2014;15:269.
Next generation sequencing based technologies are being extensively used to study transcriptomes. Among these, cap analysis of gene expression (CAGE) is specialized in detecting the most 5’ ends of RNA molecules. After mapping the sequenced reads back to a reference genome CAGE data highlights the transcriptional start sites (TSSs) and their usage at a single nucleotide resolution.
We propose a pipeline to group the single nucleotide TSS into larger reproducible peaks and compare their usage across biological states. Importantly, our pipeline discovers broad peaks as well as the fine structure of individual transcriptional start sites embedded within them. We assess the performance of our approach on a large CAGE datasets including 156 primary cell types and two cell lines with biological replicas. We demonstrate that genes have complicated structures of transcription initiation events. In particular, we discover that narrow peaks embedded in broader regions of transcriptional activity can be differentially used even if the larger region is not.
By examining the reproducible fine scaled organization of TSS we can detect many differentially regulated peaks undetected by previous approaches.
PMCID: PMC4029093  PMID: 24779366
CAGE; Peak finding; Reproducibility; Hierarchical stability
3.  ngs.plot: Quick mining and visualization of next-generation sequencing data by integrating genomic databases 
BMC Genomics  2014;15:284.
Understanding the relationship between the millions of functional DNA elements and their protein regulators, and how they work in conjunction to manifest diverse phenotypes, is key to advancing our understanding of the mammalian genome. Next-generation sequencing technology is now used widely to probe these protein-DNA interactions and to profile gene expression at a genome-wide scale. As the cost of DNA sequencing continues to fall, the interpretation of the ever increasing amount of data generated represents a considerable challenge.
We have developed ngs.plot – a standalone program to visualize enrichment patterns of DNA-interacting proteins at functionally important regions based on next-generation sequencing data. We demonstrate that ngs.plot is not only efficient but also scalable. We use a few examples to demonstrate that ngs.plot is easy to use and yet very powerful to generate figures that are publication ready.
We conclude that ngs.plot is a useful tool to help fill the gap between massive datasets and genomic information in this era of big sequencing data.
PMCID: PMC4028082  PMID: 24735413
Next-generation sequencing; Visualization; Epigenomics; Data mining; Genomic databases
4.  HGCS: an online tool for prioritizing disease-causing gene variants by biological distance 
BMC Genomics  2014;15:256.
Identifying the genotypes underlying human disease phenotypes is a fundamental step in human genetics and medicine. High-throughput genomic technologies provide thousands of genetic variants per individual. The causal genes of a specific phenotype are usually expected to be functionally close to each other. According to this hypothesis, candidate genes are picked from high-throughput data on the basis of their biological proximity to core genes — genes already known to be responsible for the phenotype. There is currently no effective gene-centric online interface for this purpose.
We describe here the human gene connectome server (HGCS), a powerful, easy-to-use interactive online tool enabling researchers to prioritize any list of genes according to their biological proximity to core genes associated with the phenotype of interest. We also make available an updated and extended version for all human gene-specific connectomes. The HGCS is freely available to noncommercial users from:
The HGCS should help investigators from diverse fields to identify new disease-causing candidate genes more effectively, via a user-friendly online interface.
PMCID: PMC4051124  PMID: 24694260
5.  seqCNA: an R package for DNA copy number analysis in cancer using high-throughput sequencing 
BMC Genomics  2014;15:178.
Deviations in the amount of genomic content that arise during tumorigenesis, called copy number alterations, are structural rearrangements that can critically affect gene expression patterns. Additionally, copy number alteration profiles allow insight into cancer discrimination, progression and complexity. On data obtained from high-throughput sequencing, improving quality through GC bias correction and keeping false positives to a minimum help build reliable copy number alteration profiles.
We introduce seqCNA, a parallelized R package for an integral copy number analysis of high-throughput sequencing cancer data. The package includes novel methodology on (i) filtering, reducing false positives, and (ii) GC content correction, improving copy number profile quality, especially under great read coverage and high correlation between GC content and copy number. Adequate analysis steps are automatically chosen based on availability of paired-end mapping, matched normal samples and genome annotation.
seqCNA, available through Bioconductor, provides accurate copy number predictions in tumoural data, thanks to the extensive filtering and better GC bias correction, while providing an integrated and parallelized workflow.
PMCID: PMC4022175  PMID: 24597965
High-throughput sequencing; Cancer; Copy number; R; Bioconductor
6.  eRNA: a graphic user interface-based tool optimized for large data analysis from high-throughput RNA sequencing 
BMC Genomics  2014;15:176.
RNA sequencing (RNA-seq) is emerging as a critical approach in biological research. However, its high-throughput advantage is significantly limited by the capacity of bioinformatics tools. The research community urgently needs user-friendly tools to efficiently analyze the complicated data generated by high throughput sequencers.
We developed a standalone tool with graphic user interface (GUI)-based analytic modules, known as eRNA. The capacity of performing parallel processing and sample management facilitates large data analyses by maximizing hardware usage and freeing users from tediously handling sequencing data. The module miRNA identification” includes GUIs for raw data reading, adapter removal, sequence alignment, and read counting. The module “mRNA identification” includes GUIs for reference sequences, genome mapping, transcript assembling, and differential expression. The module “Target screening” provides expression profiling analyses and graphic visualization. The module “Self-testing” offers the directory setups, sample management, and a check for third-party package dependency. Integration of other GUIs including Bowtie, miRDeep2, and miRspring extend the program’s functionality.
eRNA focuses on the common tools required for the mapping and quantification analysis of miRNA-seq and mRNA-seq data. The software package provides an additional choice for scientists who require a user-friendly computing environment and high-throughput capacity for large data analysis. eRNA is available for free download at
PMCID: PMC4029068  PMID: 24593312
RNA sequencing; Bioinformatics tool; Graphic user interface; Parallel processing
7.  SNPhylo: a pipeline to construct a phylogenetic tree from huge SNP data 
BMC Genomics  2014;15:162.
Phylogenetic trees are widely used for genetic and evolutionary studies in various organisms. Advanced sequencing technology has dramatically enriched data available for constructing phylogenetic trees based on single nucleotide polymorphisms (SNPs). However, massive SNP data makes it difficult to perform reliable analysis, and there has been no ready-to-use pipeline to generate phylogenetic trees from these data.
We developed a new pipeline, SNPhylo, to construct phylogenetic trees based on large SNP datasets. The pipeline may enable users to construct a phylogenetic tree from three representative SNP data file formats. In addition, in order to increase reliability of a tree, the pipeline has steps such as removing low quality data and considering linkage disequilibrium. A maximum likelihood method for the inference of phylogeny is also adopted in generation of a tree in our pipeline.
Using SNPhylo, users can easily produce a reliable phylogenetic tree from a large SNP data file. Thus, this pipeline can help a researcher focus more on interpretation of the results of analysis of voluminous data sets, rather than manipulations necessary to accomplish the analysis.
PMCID: PMC3945939  PMID: 24571581
Polymorphisms; Linkage disequilibrium; Maximum likelihood
8.  SHEAR: sample heterogeneity estimation and assembly by reference 
BMC Genomics  2014;15:84.
Personal genome assembly is a critical process when studying tumor genomes and other highly divergent sequences. The accuracy of downstream analyses, such as RNA-seq and ChIP-seq, can be greatly enhanced by using personal genomic sequences rather than standard references. Unfortunately, reads sequenced from these types of samples often have a heterogeneous mix of various subpopulations with different variants, making assembly extremely difficult using existing assembly tools. To address these challenges, we developed SHEAR (Sample Heterogeneity Estimation and Assembly by Reference;, a tool that predicts SVs, accounts for heterogeneous variants by estimating their representative percentages, and generates personal genomic sequences to be used for downstream analysis.
By making use of structural variant detection algorithms, SHEAR offers improved performance in the form of a stronger ability to handle difficult structural variant types and better computational efficiency. We compare against the lead competing approach using a variety of simulated scenarios as well as real tumor cell line data with known heterogeneous variants. SHEAR is shown to successfully estimate heterogeneity percentages in both cases, and demonstrates an improved efficiency and better ability to handle tandem duplications.
SHEAR allows for accurate and efficient SV detection and personal genomic sequence generation. It is also able to account for heterogeneous sequencing samples, such as from tumor tissue, by estimating the subpopulation percentage for each heterogeneous variant.
PMCID: PMC4007568  PMID: 24476358
Genomics; Next-generation sequencing; Sequence analysis; Assembly; Personal genome; Heterogeneity; Structural variation; Prostate cancer
9.  CodaChrome: a tool for the visualization of proteome conservation across all fully sequenced bacterial genomes 
BMC Genomics  2014;15:65.
The relationships between bacterial genomes are complicated by rampant horizontal gene transfer, varied selection pressures, acquisition of new genes, loss of genes, and divergence of genes, even in closely related lineages. As more and more bacterial genomes are sequenced, organizing and interpreting the incredible amount of relational information that connects them becomes increasingly difficult.
We have developed CodaChrome (, a one-versus-all proteome comparison tool that allows the user to visually investigate the relationship between a bacterial proteome of interest and the proteomes encoded by every other bacterial genome recorded in GenBank in a massive interactive heat map. This tool has allowed us to rapidly identify the most highly conserved proteins encoded in the bacterial pan-genome, fast-clock genes useful for subtyping of bacterial species, the evolutionary history of an indel in the Sphingobium lineage, and an example of horizontal gene transfer from a member of the genus Enterococcus to a recent ancestor of Helicobacter pylori.
CodaChrome is a user-friendly and powerful tool for simultaneously visualizing relationships between thousands of proteomes.
PMCID: PMC3908345  PMID: 24460813
10.  MafFilter: a highly flexible and extensible multiple genome alignment files processor 
BMC Genomics  2014;15:53.
Sequence alignments are the starting point for most evolutionary and comparative analyses. Full genome sequences can be compared to study patterns of within and between species variation. Genome sequence alignments are complex structures containing information such as coordinates, quality scores and synteny structure, which are stored in Multiple Alignment Format (MAF) files. Processing these alignments therefore involves parsing and manipulating typically large MAF files in an efficient way.
MafFilter is a command-line driven program written in C++ that enables the processing of genome alignments stored in the Multiple Alignment Format in an efficient and extensible manner. It provides an extensive set of tools which can be parametrized and combined by the user via option files. We demonstrate the software’s functionality and performance on several biological examples covering Primate genomics and fungal population genomics. Example analyses involve window-based alignment filtering, feature extractions and various statistics, phylogenetics and population genomics calculations.
MafFilter is a highly efficient and flexible tool to analyse multiple genome alignments. By allowing the user to combine a large set of available methods, as well as designing his/her own, it enables the design of custom data filtering and analysis pipelines for genomic studies. MafFilter is an open source software available at
PMCID: PMC3904536  PMID: 24447531
11.  Using a color-coded ambigraphic nucleic acid notation to visualize conserved palindromic motifs within and across genomes 
BMC Genomics  2014;15:52.
Ambiscript is a graphically-designed nucleic acid notation that uses symbol symmetries to support sequence complementation, highlight biologically-relevant palindromes, and facilitate the analysis of consensus sequences. Although the original Ambiscript notation was designed to easily represent consensus sequences for multiple sequence alignments, the notation’s black-on-white ambiguity characters are unable to reflect the statistical distribution of nucleotides found at each position. We now propose a color-augmented ambigraphic notation to encode the frequency of positional polymorphisms in these consensus sequences.
We have implemented this color-coding approach by creating an Adobe Flash® application ( that shades and colors modified Ambiscript characters according to the prevalence of the encoded nucleotide at each position in the alignment. The resulting graphic helps viewers perceive biologically-relevant patterns in multiple sequence alignments by uniquely combining color, shading, and character symmetries to highlight palindromes and inverted repeats in conserved DNA motifs.
Juxtaposing an intuitive color scheme over the deliberate character symmetries of an ambigraphic nucleic acid notation yields a highly-functional nucleic acid notation that maximizes information content and successfully embodies key principles of graphic excellence put forth by the statistician and graphic design theorist, Edward Tufte.
PMCID: PMC3916809  PMID: 24447494
Notation; Ambigram; Palindrome; Motif; Nucleotide; Color-coded
12.  ITEP: An integrated toolkit for exploration of microbial pan-genomes 
BMC Genomics  2014;15:8.
Comparative genomics is a powerful approach for studying variation in physiological traits as well as the evolution and ecology of microorganisms. Recent technological advances have enabled sequencing large numbers of related genomes in a single project, requiring computational tools for their integrated analysis. In particular, accurate annotations and identification of gene presence and absence are critical for understanding and modeling the cellular physiology of newly sequenced genomes. Although many tools are available to compare the gene contents of related genomes, new tools are necessary to enable close examination and curation of protein families from large numbers of closely related organisms, to integrate curation with the analysis of gain and loss, and to generate metabolic networks linking the annotations to observed phenotypes.
We have developed ITEP, an Integrated Toolkit for Exploration of microbial Pan-genomes, to curate protein families, compute similarities to externally-defined domains, analyze gene gain and loss, and generate draft metabolic networks from one or more curated reference network reconstructions in groups of related microbial species among which the combination of core and variable genes constitute the their "pan-genomes". The ITEP toolkit consists of: (1) a series of modular command-line scripts for identification, comparison, curation, and analysis of protein families and their distribution across many genomes; (2) a set of Python libraries for programmatic access to the same data; and (3) pre-packaged scripts to perform common analysis workflows on a collection of genomes. ITEP’s capabilities include de novo protein family prediction, ortholog detection, analysis of functional domains, identification of core and variable genes and gene regions, sequence alignments and tree generation, annotation curation, and the integration of cross-genome analysis and metabolic networks for study of metabolic network evolution.
ITEP is a powerful, flexible toolkit for generation and curation of protein families. ITEP's modular design allows for straightforward extension as analysis methods and tools evolve. By integrating comparative genomics with the development of draft metabolic networks, ITEP harnesses the power of comparative genomics to build confidence in links between genotype and phenotype and helps disambiguate gene annotations when they are evaluated in both evolutionary and metabolic network contexts.
PMCID: PMC3890548  PMID: 24387194
Comparative genomics; Clustering; Curation; Database; Metabolic networks; Orthologs; Pan-genome; Phylogenetics
13.  PSP: rapid identification of orthologous coding genes under positive selection across multiple closely related prokaryotic genomes 
BMC Genomics  2013;14:924.
With genomic sequences of many closely related bacterial strains made available by deep sequencing, it is now possible to investigate trends in prokaryotic microevolution. Positive selection is a sub-process of microevolution, in which a particular mutation is favored, causing the allele frequency to continuously shift in one direction. Wide scanning of prokaryotic genomes has shown that positive selection at the molecular level is much more frequent than expected. Genes with significant positive selection may play key roles in bacterial adaption to different environmental pressures. However, selection pressure analyses are computationally intensive and awkward to configure.
Here we describe an open access web server, which is designated as PSP (Positive Selection analysis for Prokaryotic genomes) for performing evolutionary analysis on orthologous coding genes, specially designed for rapid comparison of dozens of closely related prokaryotic genomes. Remarkably, PSP facilitates functional exploration at the multiple levels by assignments and enrichments of KO, GO or COG terms. To illustrate this user-friendly tool, we analyzed Escherichia coli and Bacillus cereus genomes and found that several genes, which play key roles in human infection and antibiotic resistance, show significant evidence of positive selection. PSP is freely available to all users without any login requirement at:
PSP ultimately allows researchers to do genome-scale analysis for evolutionary selection across multiple prokaryotic genomes rapidly and easily, and identify the genes undergoing positive selection, which may play key roles in the interactions of host-pathogen and/or environmental adaptation.
PMCID: PMC3882776  PMID: 24373418
Orthologous genes; Positive selection; Synonymous and nonsynonymous substitutions; Bacterial microevolution; Bacillus cereus; Escherichia coli
14.  TIPMaP: a web server to establish transcript isoform profiles from reliable microarray probes 
BMC Genomics  2013;14:922.
Standard 3′ Affymetrix gene expression arrays have contributed a significantly higher volume of existing gene expression data than other microarray platforms. These arrays were designed to identify differentially expressed genes, but not their alternatively spliced transcript forms. No resource can currently identify expression pattern of specific mRNA forms using these microarray data, even though it is possible to do this.
We report a web server for expression profiling of alternatively spliced transcripts using microarray data sets from 31 standard 3′ Affymetrix arrays for human, mouse and rat species. The tool has been experimentally validated for mRNAs transcribed or not-detected in a human disease condition (non-obstructive azoospermia, a male infertility condition). About 4000 gene expression datasets were downloaded from a public repository. ‘Good probes’ with complete coverage and identity to latest reference transcript sequences were first identified. Using them, ‘Transcript specific probe-clusters’ were derived for each platform and used to identify expression status of possible transcripts. The web server can lead the user to datasets corresponding to specific tissues, conditions via identifiers of the microarray studies or hybridizations, keywords, official gene symbols or reference transcript identifiers. It can identify, in the tissues and conditions of interest, about 40% of known transcripts as ‘transcribed’, ‘not-detected’ or ‘differentially regulated’. Corresponding additional information for probes, genes, transcripts and proteins can be viewed too. We identified the expression of transcripts in a specific clinical condition and validated a few of these transcripts by experiments (using reverse transcription followed by polymerase chain reaction). The experimental observations indicated higher agreements with the web server results, than contradictions. The tool is accessible at
The newly developed online tool forms a reliable means for identification of alternatively spliced transcript-isoforms that may be differentially expressed in various tissues, cell types or physiological conditions. Thus, by making better use of existing data, TIPMaP avoids the dependence on precious tissue-samples, in experiments with a goal to establish expression profiles of alternative splice forms – at least in some cases.
PMCID: PMC3884118  PMID: 24373374
Alternative splicing; Alternatively spliced; Microarray; Affymetrix; Azoospermia; Transcript isoforms; mRNA isoforms; Transcriptome; Gene expression
15.  POMO - Plotting Omics analysis results for Multiple Organisms 
BMC Genomics  2013;14:918.
Systems biology experiments studying different topics and organisms produce thousands of data values across different types of genomic data. Further, data mining analyses are yielding ranked and heterogeneous results and association networks distributed over the entire genome. The visualization of these results is often difficult and standalone web tools allowing for custom inputs and dynamic filtering are limited.
We have developed POMO (, an interactive web-based application to visually explore omics data analysis results and associations in circular, network and grid views. The circular graph represents the chromosome lengths as perimeter segments, as a reference outer ring, such as cytoband for human. The inner arcs between nodes represent the uploaded network. Further, multiple annotation rings, for example depiction of gene copy number changes, can be uploaded as text files and represented as bar, histogram or heatmap rings. POMO has built-in references for human, mouse, nematode, fly, yeast, zebrafish, rice, tomato, Arabidopsis, and Escherichia coli. In addition, POMO provides custom options that allow integrated plotting of unsupported strains or closely related species associations, such as human and mouse orthologs or two yeast wild types, studied together within a single analysis. The web application also supports interactive label and weight filtering. Every iterative filtered result in POMO can be exported as image file and text file for sharing or direct future input.
The POMO web application is a unique tool for omics data analysis, which can be used to visualize and filter the genome-wide networks in the context of chromosomal locations as well as multiple network layouts. With the several illustration and filtering options the tool supports the analysis and visualization of any heterogeneous omics data analysis association results for many organisms. POMO is freely available and does not require any installation or registration.
PMCID: PMC3880012  PMID: 24365393
Omics; Association; Visualization; Ortholog; Phenolog; Genome-wide; Network; Model organism
16.  BS-Seeker2: a versatile aligning pipeline for bisulfite sequencing data 
BMC Genomics  2013;14:774.
DNA methylation is an important epigenetic modification involved in many biological processes. Bisulfite treatment coupled with high-throughput sequencing provides an effective approach for studying genome-wide DNA methylation at base resolution. Libraries such as whole genome bisulfite sequencing (WGBS) and reduced represented bisulfite sequencing (RRBS) are widely used for generating DNA methylomes, demanding efficient and versatile tools for aligning bisulfite sequencing data.
We have developed BS-Seeker2, an updated version of BS Seeker, as a full pipeline for mapping bisulfite sequencing data and generating DNA methylomes. BS-Seeker2 improves mappability over existing aligners by using local alignment. It can also map reads from RRBS library by building special indexes with improved efficiency and accuracy. Moreover, BS-Seeker2 provides additional function for filtering out reads with incomplete bisulfite conversion, which is useful in minimizing the overestimation of DNA methylation levels. We also defined CGmap and ATCGmap file formats for full representations of DNA methylomes, as part of the outputs of BS-Seeker2 pipeline together with BAM and WIG files.
Our evaluations on the performance show that BS-Seeker2 works efficiently and accurately for both WGBS data and RRBS data. BS-Seeker2 is freely available at and the Galaxy server.
PMCID: PMC3840619  PMID: 24206606
DNA methylation; Bisulfite sequencing aligner; WGBS; RRBS; BS Seeker; Bisulfite conversion failure; Galaxy toolshed
17.  Guide: a desktop application for analysing gene expression data 
BMC Genomics  2013;14:688.
Multiplecompeting bioinformatics tools exist for next-generation sequencing data analysis. Many of these tools are available as R/Bioconductor modules, and it can be challenging for the bench biologist without any programming background to quickly analyse genomics data. Here, we present an application that is designed to be simple to use, while leveraging the power of R as the analysis engine behind the scenes.
Genome Informatics Data Explorer (Guide) is a desktop application designed for the bench biologist to analyse RNA-seq and microarray gene expression data. It requires a text file of summarised read counts or expression values as input data, and performs differential expression analyses at both the gene and pathway level. It uses well-established R/Bioconductor packages such as limma for its analyses, without requiring the user to have specific knowledge of the underlying R functions. Results are presented in figures or interactive tables which integrate useful data from multiple sources such as gene annotation and orthologue data. Advanced options include the ability to edit R commands to customise the analysis pipeline.
Guide is a desktop application designed to query gene expression data in a user-friendly way while automatically communicating with R. Its customisation options make it possible to use different bioinformatics tools available through R/Bioconductor for its analyses, while keeping the core usage simple. Guide is written in the cross-platform framework of Qt, and is freely available for use from
PMCID: PMC3815230  PMID: 24093424
Data analysis; R; Gene expression; RNA-seq; Microarray; Differential expression; Software
18.  InsertionMapper: a pipeline tool for the identification of targeted sequences from multidimensional high throughput sequencing data 
BMC Genomics  2013;14:679.
The advent of next-generation high-throughput technologies has revolutionized whole genome sequencing, yet some experiments require sequencing only of targeted regions of the genome from a very large number of samples. These regions can be amplified by PCR and sequenced by next-generation methods using a multidimensional pooling strategy. However, there is at present no available generalized tool for the computational analysis of target-enriched NGS data from multidimensional pools.
Here we present InsertionMapper, a pipeline tool for the identification of targeted sequences from multidimensional high throughput sequencing data. InsertionMapper consists of four independently working modules: Data Preprocessing, Database Modeling, Dimension Deconvolution and Element Mapping. We illustrate InsertionMapper with an example from our project 'New reverse genetics resources for maize’, which aims to sequence-index a collection of 15,000 independent insertion sites of the transposon Ds in maize. Identified sequences are validated by PCR assays. This pipeline tool is applicable to similar scenarios requiring analysis of the tremendous output of short reads produced in NGS sequencing experiments of targeted genome sequences.
InsertionMapper is proven efficacious for the identification of target-enriched sequences from multidimensional high throughput sequencing data. With adjustable parameters and experiment configurations, this tool can save great computational effort to biologists interested in identifying their sequences of interest within the huge output of modern DNA sequencers. InsertionMapper is freely accessible at and
PMCID: PMC3850689  PMID: 24090499
Next-generation sequencing; Sequence identification; Target enrichment; Multidimensional pooling
19.  Predicting whole genome protein interaction networks from primary sequence data in model and non-model organisms using ENTS 
BMC Genomics  2013;14:608.
The large-scale identification of physical protein-protein interactions (PPIs) is an important step toward understanding how biological networks evolve and generate emergent phenotypes. However, experimental identification of PPIs is a laborious and error-prone process, and current methods of PPI prediction tend to be highly conservative or require large amounts of functional data that may not be available for newly-sequenced organisms.
In this study we demonstrate a random-forest based technique, ENTS, for the computational prediction of protein-protein interactions based only on primary sequence data. Our approach is able to efficiently predict interactions on a whole-genome scale for any eukaryotic organism, using pairwise combinations of conserved domains and predicted subcellular localization of proteins as input features. We present the first predicted interactome for the forest tree Populus trichocarpa in addition to the predicted interactomes for Saccharomyces cerevisiae, Homo sapiens, Mus musculus, and Arabidopsis thaliana. Comparing our approach to other PPI predictors, we find that ENTS performs comparably to or better than a number of existing approaches, including several that utilize a variety of functional information for their predictions. We also find that the predicted interactions are biologically meaningful, as indicated by similarity in functional annotations and enrichment of co-expressed genes in public microarray datasets. Furthermore, we demonstrate some of the biological insights that can be gained from these predicted interaction networks. We show that the predicted interactions yield informative groupings of P. trichocarpa metabolic pathways, literature-supported associations among human disease states, and theory-supported insight into the evolutionary dynamics of duplicated genes in paleopolyploid plants.
We conclude that the ENTS classifier will be a valuable tool for the de novo annotation of genome sequences, providing initial clues about regulatory and metabolic network topology, and revealing relationships that are not immediately obvious from traditional homology-based annotations.
PMCID: PMC3848842  PMID: 24015873
20.  Confero: an integrated contrast data and gene set platform for computational analysis and biological interpretation of omics data 
BMC Genomics  2013;14:514.
High-throughput omics technologies such as microarrays and next-generation sequencing (NGS) have become indispensable tools in biological research. Computational analysis and biological interpretation of omics data can pose significant challenges due to a number of factors, in particular the systems integration required to fully exploit and compare data from different studies and/or technology platforms. In transcriptomics, the identification of differentially expressed genes when studying effect(s) or contrast(s) of interest constitutes the starting point for further downstream computational analysis (e.g. gene over-representation/enrichment analysis, reverse engineering) leading to mechanistic insights. Therefore, it is important to systematically store the full list of genes with their associated statistical analysis results (differential expression, t-statistics, p-value) corresponding to one or more effect(s) or contrast(s) of interest (shortly termed as ” contrast data”) in a comparable manner and extract gene sets in order to efficiently support downstream analyses and further leverage data on a long-term basis. Filling this gap would open new research perspectives for biologists to discover disease-related biomarkers and to support the understanding of molecular mechanisms underlying specific biological perturbation effects (e.g. disease, genetic, environmental, etc.).
To address these challenges, we developed Confero, a contrast data and gene set platform for downstream analysis and biological interpretation of omics data. The Confero software platform provides storage of contrast data in a simple and standard format, data transformation to enable cross-study and platform data comparison, and automatic extraction and storage of gene sets to build new a priori knowledge which is leveraged by integrated and extensible downstream computational analysis tools. Gene Set Enrichment Analysis (GSEA) and Over-Representation Analysis (ORA) are currently integrated as an analysis module as well as additional tools to support biological interpretation. Confero is a standalone system that also integrates with Galaxy, an open-source workflow management and data integration system. To illustrate Confero platform functionality we walk through major aspects of the Confero workflow and results using the Bioconductor estrogen package dataset.
Confero provides a unique and flexible platform to support downstream computational analysis facilitating biological interpretation. The system has been designed in order to provide the researcher with a simple, innovative, and extensible solution to store and exploit analyzed data in a sustainable and reproducible manner thereby accelerating knowledge-driven research. Confero source code is freely available from
PMCID: PMC3750322  PMID: 23895370
Gene expression; Contrast data; Gene set; Gene set enrichment; Omics; Microarray; Next-generation sequencing; Reproducible research system; Knowledge acquisition
21.  Cloud-based uniform ChIP-Seq processing tools for modENCODE and ENCODE 
BMC Genomics  2013;14:494.
Funded by the National Institutes of Health (NIH), the aim of the Model Organism ENCyclopedia of DNA Elements (modENCODE) project is to provide the biological research community with a comprehensive encyclopedia of functional genomic elements for both model organisms C. elegans (worm) and D. melanogaster (fly). With a total size of just under 10 terabytes of data collected and released to the public, one of the challenges faced by researchers is to extract biologically meaningful knowledge from this large data set. While the basic quality control, pre-processing, and analysis of the data has already been performed by members of the modENCODE consortium, many researchers will wish to reinterpret the data set using modifications and enhancements of the original protocols, or combine modENCODE data with other data sets. Unfortunately this can be a time consuming and logistically challenging proposition.
In recognition of this challenge, the modENCODE DCC has released uniform computing resources for analyzing modENCODE data on Galaxy (, on the public Amazon Cloud (, and on the private Bionimbus Cloud for genomic research ( In particular, we have released Galaxy workflows for interpreting ChIP-seq data which use the same quality control (QC) and peak calling standards adopted by the modENCODE and ENCODE communities. For convenience of use, we have created Amazon and Bionimbus Cloud machine images containing Galaxy along with all the modENCODE data, software and other dependencies.
Using these resources provides a framework for running consistent and reproducible analyses on modENCODE data, ultimately allowing researchers to use more of their time using modENCODE data, and less time moving it around.
PMCID: PMC3734164  PMID: 23875683
22.  Rainbow: a tool for large-scale whole-genome sequencing data analysis using cloud computing 
BMC Genomics  2013;14:425.
Technical improvements have decreased sequencing costs and, as a result, the size and number of genomic datasets have increased rapidly. Because of the lower cost, large amounts of sequence data are now being produced by small to midsize research groups. Crossbow is a software tool that can detect single nucleotide polymorphisms (SNPs) in whole-genome sequencing (WGS) data from a single subject; however, Crossbow has a number of limitations when applied to multiple subjects from large-scale WGS projects. The data storage and CPU resources that are required for large-scale whole genome sequencing data analyses are too large for many core facilities and individual laboratories to provide. To help meet these challenges, we have developed Rainbow, a cloud-based software package that can assist in the automation of large-scale WGS data analyses.
Here, we evaluated the performance of Rainbow by analyzing 44 different whole-genome-sequenced subjects. Rainbow has the capacity to process genomic data from more than 500 subjects in two weeks using cloud computing provided by the Amazon Web Service. The time includes the import and export of the data using Amazon Import/Export service. The average cost of processing a single sample in the cloud was less than 120 US dollars. Compared with Crossbow, the main improvements incorporated into Rainbow include the ability: (1) to handle BAM as well as FASTQ input files; (2) to split large sequence files for better load balance downstream; (3) to log the running metrics in data processing and monitoring multiple Amazon Elastic Compute Cloud (EC2) instances; and (4) to merge SOAPsnp outputs for multiple individuals into a single file to facilitate downstream genome-wide association studies.
Rainbow is a scalable, cost-effective, and open-source tool for large-scale WGS data analysis. For human WGS data sequenced by either the Illumina HiSeq 2000 or HiSeq 2500 platforms, Rainbow can be used straight out of the box. Rainbow is available for third-party implementation and use, and can be downloaded from
PMCID: PMC3698007  PMID: 23802613
Cloud computing; Whole genome sequencing; Single nucleotide polymorphism; SNP; Next generation sequencing; Software
23.  Web-based visual analysis for high-throughput genomics 
BMC Genomics  2013;14:397.
Visualization plays an essential role in genomics research by making it possible to observe correlations and trends in large datasets as well as communicate findings to others. Visual analysis, which combines visualization with analysis tools to enable seamless use of both approaches for scientific investigation, offers a powerful method for performing complex genomic analyses. However, there are numerous challenges that arise when creating rich, interactive Web-based visualizations/visual analysis applications for high-throughput genomics. These challenges include managing data flow from Web server to Web browser, integrating analysis tools and visualizations, and sharing visualizations with colleagues.
We have created a platform simplifies the creation of Web-based visualization/visual analysis applications for high-throughput genomics. This platform provides components that make it simple to efficiently query very large datasets, draw common representations of genomic data, integrate with analysis tools, and share or publish fully interactive visualizations. Using this platform, we have created a Circos-style genome-wide viewer, a generic scatter plot for correlation analysis, an interactive phylogenetic tree, a scalable genome browser for next-generation sequencing data, and an application for systematically exploring tool parameter spaces to find good parameter values. All visualizations are interactive and fully customizable. The platform is integrated with the Galaxy ( genomics workbench, making it easy to integrate new visual applications into Galaxy.
Visualization and visual analysis play an important role in high-throughput genomics experiments, and approaches are needed to make it easier to create applications for these activities. Our framework provides a foundation for creating Web-based visualizations and integrating them into Galaxy. Finally, the visualizations we have created using the framework are useful tools for high-throughput genomics experiments.
PMCID: PMC3691752  PMID: 23758618
Visualization; Visual analysis; Galaxy; Genome browser; Circos; Phylogenetic tree
24.  Unsupervised genome-wide recognition of local relationship patterns 
BMC Genomics  2013;14:347.
Phenomena such as incomplete lineage sorting, horizontal gene transfer, gene duplication and subsequent sub- and neo-functionalisation can result in distinct local phylogenetic relationships that are discordant with species phylogeny. In order to assess the possible biological roles for these subdivisions, they must first be identified and characterised, preferably on a large scale and in an automated fashion.
We developed Saguaro, a combination of a Hidden Markov Model (HMM) and a Self Organising Map (SOM), to characterise local phylogenetic relationships among aligned sequences using cacti, matrices of pair-wise distance measures. While the HMM determines the genomic boundaries from aligned sequences, the SOM hypothesises new cacti in an unsupervised and iterative fashion based on the regions that were modelled least well by existing cacti. After testing the software on simulated data, we demonstrate the utility of Saguaro by testing two different data sets: (i) 181 Dengue virus strains, and (ii) 5 primate genomes. Saguaro identifies regions under lineage-specific constraint for the first set, and genomic segments that we attribute to incomplete lineage sorting in the second dataset. Intriguingly for the primate data, Saguaro also classified an additional ~3% of the genome as most incompatible with the expected species phylogeny. A substantial fraction of these regions was found to overlap genes associated with both the innate and adaptive immune systems.
Saguaro detects distinct cacti describing local phylogenetic relationships without requiring any a priori hypotheses. We have successfully demonstrated Saguaro’s utility with two contrasting data sets, one containing many members with short sequences (Dengue viral strains: n = 181, genome size = 10,700 nt), and the other with few members but complex genomes (related primate species: n = 5, genome size = 3 Gb), suggesting that the software is applicable to a wide variety of experimental populations. Saguaro is written in C++, runs on the Linux operating system, and can be downloaded from
PMCID: PMC3669000  PMID: 23706020
25.  ContigScape: a Cytoscape plugin facilitating microbial genome gap closing 
BMC Genomics  2013;14:289.
With the emergence of next-generation sequencing, the availability of prokaryotic genome sequences is expanding rapidly. A total of 5,276 genomes have been released since 2008, yet only 1,692 genomes were complete. The final phase of microbial genome sequencing, particularly gap closing, is frequently the rate-limiting step either because of complex genomic structures that cause sequence bias even with high genomic coverage, or the presence of repeat sequences that may cause gaps in assembly.
We have developed a Cytoscape plugin to facilitate gap closing for high-throughput sequencing data from microbial genomes. This plugin is capable of interactively displaying the relationships among genomic contigs derived from various sequencing formats. The sequence contigs of plasmids and special repeats (IS elements, ribosomal RNAs, terminal repeats, etc.) can be displayed as well.
Displaying relationships between contigs using graphs in Cytoscape rather than tables provides a more straightforward visual representation. This will facilitate a faster and more precise determination of the linkages among contigs and greatly improve the efficiency of gap closing.
PMCID: PMC3651407  PMID: 23627759
ContigScape; Repeat contig; Microbial; Visualization; Linkage; Gap closing

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