Abstract
The catfish genome database, cBARBEL (abbreviated from catfish Breeder And Researcher Bioinformatics Entry Location) is an online open-access database for genome biology of ictalurid catfish (Ictalurus spp.). It serves as a comprehensive, integrative platform for all aspects of catfish genetics, genomics and related data resources. cBARBEL provides BLAST-based, fuzzy and specific search functions, visualization of catfish linkage, physical and integrated maps, a catfish EST contig viewer with SNP information overlay, and GBrowse-based organization of catfish genomic data based on sequence similarity with zebrafish chromosomes. Subsections of the database are tightly related, allowing a user with a sequence or search string of interest to navigate seamlessly from one area to another. As catfish genome sequencing proceeds and ongoing quantitative trait loci (QTL) projects bear fruit, cBARBEL will allow rapid data integration and dissemination within the catfish research community and to interested stakeholders. cBARBEL can be accessed at http://catfishgenome.org.
INTRODUCTION
Catfish (Ictalurus spp.) is an important aquaculture species in the United States, accounting for >60% of domestic aquaculture production. While channel catfish (Ictalurus punctatus) accounts for the large majority of farm-raised catfish, increasing numbers of channel catfish female × blue catfish (Ictalurus furcatus) male hybrids are being cultured. Facing rising feed costs and stiff international competition, catfish producers require improvement in fish production and performance traits such as disease resistance, growth rate and feed conversion efficiency to maintain profitability. Efficient utilization of the natural diversity of trait phenotypes already present in different species, strains and hybrids of catfish for selection of superior broodstock, requires the identification of genetic underpinnings of trait differences. Toward this end and eventual marker-assisted selection (MAS), significant genome resources have been developed in catfish. These include over a half million expressed sequence tags (ESTs) (1–6), a large number of genome sequences generated from bacterial artificial chromosome (BAC) ends (7), genetic linkage maps (8–10), genome physical maps (11,12), tens of thousands of microsatellite markers (13,14), hundreds of thousands of single-nucleotide polymorphisms (SNPs) (15), over 10 000 full-length cDNAs (flcDNA; 16) and an alternative splicing database (17). Additionally, USDA NIFA funding has been secured to allow catfish whole-genome sequencing and the development of high-density SNP chips for catfish. With these and many more genome-oriented projects from catfish currently underway, the catfish research community needed a central repository for storing and integrating genomic data and a bioinformatic entry location for public access to currently inaccessible specialized data sets. To meet this need, we have created a catfish genome database, cBARBEL, the Catfish Breeder and Researcher Bioinformatic Entry Location, a title that makes use of the distinctive whisker-like organs that give catfish their name.
cBARBEL represents one of the first comprehensive bioinformatic databases for an aquaculture species, although genome sequencing is planned or proceeding in close to a dozen different species from this fast-growing sector. cBARBEL provides wide-ranging query functions to facilitate user access to a host of catfish genome resources and integrates a variety of previously scattered data types. Here, we present an overview of cBARBEL search tools, platforms and functions connecting catfish EST, fl-cDNA, SNP, BAC-end sequence (BES), molecular marker, linkage map and physical map data. cBARBEL can be accessed at http://catfishgenome.org/.
RESULTS AND DISCUSSION
cBARBEL database schematic
The cBARBEL database schematic is shown in Figure 1, allowing visualization of potential data connections. The integration of existing and forthcoming catfish genome resources provided by the cBARBEL platform should speed research progress. Utilizing cBARBEL, for example, a user searching for a given gene in catfish is able to search with a similar sequence from a related species, such as zebrafish, identify matching catfish ESTs, identify the corresponding EST contig, identify SNP markers within that contig, visualize linkage map position of these markers and relate linkage and physical map locations through a comparative map system, all by navigating through a series of intuitive links. To do a similar search without cBARBEL would require interrogating a series of separate public and local databases, relating disparate nomenclatures, would not include visualization and could take well over 30 min for an individual query.
Figure 1.
The cBARBEL database schematic showing database components and clickable, searchable connections between catfish genome resources. BES, BAC-end sequence; SNP, single-nucleotide polymorphism; EST, expressed sequence tag.
Implementation
Several software packages were used in the construction of cBARBEL database prerequisites including: (i) the operating system, Ubuntu 9.10 Linux system (http://www.ubuntu.com/); (ii) Apache web server version 2.0 (http://www.apache.org/); (iii) MySQL database management system, version 5.1 (http://www.mysql.com/); and (iv) Selective Perl modules and configuration of Bioperl and PHP (www.perl.org; www.bioperl.org; http://php.net/). The Generic Genome Browser (GBrowse) package (v1.7), a component of the Generic Model Organism Project (GMOD), was utilized for display of the catfish physical map, EST contig viewer and zebrafish comparative GBrowse. Another GMOD component, CMap (18), was used to display and align genetic and physical (FPC) maps.
General features of cBARBEL components and tools
cBARBEL currently is organized around three components: nucleotide sequence, genetic markers and maps. These components are brought together by search tools and multi-directional links. Features of each of these components are described briefly below.
Sequence similarity searches of catfish databases
BLAST searches can be carried out to query all or a subset of catfish sequences including catfish ESTs, catfish BES, full-length cDNA and catfish all (Figure 2). Search results provide further links to NCBI records and location of the hit on the catfish physical map (BES), EST contig viewer (EST) and zebrafish GBrowse chromosomes (All).
Figure 2.
cBARBEL database sequence search functions. Searches can be conducted by BLAST against local databases of catfish ESTs, catfish BES, full-length cDNA or all. Searches can also be conducted using fuzzy or specific searches for ESTs, BES, flcDNA and genetic markers and associated subcategories. Examples for all query types are provided.
Specific search function of catfish databases
The specific search function can provide data access using a variety of search terms (Figure 2). Catfish ESTs can be queried using GenBank accession numbers, marker names for those ESTs containing a SNP or microsatellite marker, and EST contig ID. Search results provide sequence links as well as deeper connections to the EST contig viewer, zebrafish GBrowse comparative alignments and linkage map position, where appropriate. Similar searches can be carried out for BES and fl-cDNA. Those looking for a marker of interest can use fuzzy or specific search terms. For example, a search such as ‘AUEST’ returns all marker information for EST microsatellite markers generated at Auburn University. The resulting marker table contains the accession number of the relevant sequence, primer information, marker status (polymorphic, not polymorphic, untested), linkage map position, physical map position (BES) and EST-contig (EST).
Zebrafish GBrowse genomic viewer versus catfish genomic data sets
GBrowse is a GMOD tool that displays features of the genome aligned to a genomic sequence (19). GBrowse is easily customized to allow a variety of data tracks and third-party data types to be visualized. Catfish whole-genome sequencing is underway, but, in the interim, we have aligned a number of catfish genome data types to the genome sequence of zebrafish, the closest evolutionarily related species with an available sequenced genome (Figure 3). The alignments (based on tblastx similarity) help to organize catfish data on a genome scale, and harnessing synteny between the two species has proven useful in gene isolation and QTL fine-mapping studies. As catfish genome assembly proceeds, this comparative approach should also prove useful in scaffolding catfish supercontigs. By default, cBARBEL presents a view of catfish EST contigs, catfish singleton ESTs, catfish BES, and catfish fl-cDNA aligned to zebrafish chromosomal sequences (Figure 3). For each of these tracks, clicking on a feature provides a related link to the NCBI server or a link to a local copy of the sequence information. Users can locate a specific region of interest by entering a specific sequence range or by dragging the selection box to specify a chromosomal region. Additionally, cBARBEL BLAST and specific search outputs include zebrafish GBrowse links to allow connectivity to other database sectors.
Figure 3.
Mapping and organization of catfish genomic data based on homology with zebrafish chromosomal regions. In the absence of genome sequence for catfish, catfish EST contigs, singleton ESTs, BES, flcDNA were mapped based on tblastx results onto zebrafish chromosomes in the GBrowse package. Zoom features allow for examination of gene level to full chromosome level alignments. Mapped elements within each track are clickable to NCBI or local databases providing specific sequence information.
Catfish EST contig viewer
The GBrowse package was also utilized to create a catfish EST contig viewer displaying the alignment of individual ESTs on the contig consensus sequence. A SNP track was also added allowing visualization of SNP density and position within the EST contig (Figure 4). For the EST track, clicking on an individual feature provides a related link to NCBI-based sequence information. Clicking on SNP entries allows the user to navigate to further SNP information in the AutoSNP program including SNP allele frequency, type, and position. For the contig track, a link is provided to local sequence information. As with the zebrafish GBrowse view, cBARBEL BLAST and specific search outputs include links to the EST contig viewer.
Figure 4.
Catfish EST contig viewer within the GBrowse package. Tracks include Contig, EST and SNP and allow easy visualization of contig component sequences and physical position of SNPs along the consensus sequence. Each track is clickable to sequence information via NCBI (EST) or local database (contig) or AutoSNP (SNP) for detailed SNP allele frequency, type and position.
Catfish physical map based on BAC contig
We previously reported the construction of a fingerprint contig (FPC) BAC-based physical map with 3307 contigs spanning the catfish genome (12), and are currently engaged in efforts to integrate the physical map with catfish linkage maps using BES-associated microsatellites. This map was previously displayed using a Java-based program, WebFPC, which did not allow efficient search or data integration options. For example, BES associated with BAC clones could not be searched or visualized with WebFPC. To remedy these issues, we adapted GBrowse to display both BAC contigs and BES information in a searchable, integrated format (Figure 5). BAC clones are displayed using their FPC position within a given contig. Custom scripts were developed to indicate the presence of BES within a clone using blue boxes and to allow BES sequence retrieval via NCBI link by clicking on the desired clone. As with other sections, cBARBEL BLAST and specific search outputs include physical map links where appropriate.
Figure 5.
Catfish BAC-based physical map. Catfish BAC contigs are viewable and searchable in the GBrowse format. Associated BESs are indicated by blue clone ends and NCBI-based sequences can be retrieved by clicking a clone of interest.
Catfish linkage map
A linkage map of catfish has been constructed based on genotyping of EST-based microsatellites, SNP markers and BES-based microsatellite markers on backcross hybrid (channel × blue) families (10). Efforts are ongoing to increase marker density using BES-associated microsatellites and SNP markers. Marker information for the 29 linkage groups is displayed in table format with marker name and map position (cM). Clicking on marker name allows navigation to a table providing more detailed information including primer sequences, marker status and relevant EST contig (EST) or physical map (BES) locations. As additional markers are genotyped and new maps constructed, this information is rapidly updated.
Catfish CMap–map integration
Integration of catfish linkage and physical maps is ongoing, based largely on the mapping of BES microsatellites from physical map contigs onto the catfish linkage map. While details of integration can be gathered in table format in other cBARBEL sections, we used CMap to provide visualization of LG-level map integration (Figure 6). Links are provided to each of the 29 linkage groups arrayed alongside corresponding physical map contigs using predetermined settings. A link is also provided to allow users to access numerous different display settings including the choice to view graphic representations of linkage groups alone. As with the linkage maps, data is continually updated as additional markers are genotyped.
Figure 6.
CMap-based visualization of early integration of catfish linkage and physical maps through mapping of BES microsatellites. Catfish linkage group 26 (LG26) is provided as an example on the left, and BES-associated contigs are shown on the right. CMap allows numerous options for viewing and comparing the catfish linkage and physical maps.
Other tools and features
Under Tools, cBARBEL provides links to simple daily-use informatics tools that database users may need while accessing cBARBEL, including the external NCBI BLAST server, SMART domain search, ClustalW-based multiple sequence alignments and basic nucleotide to amino acid translation. These tools are provided in-frame so that the user does not need to leave the cBARBEL site for small-scale sequence analysis. Publications relevant to catfish genomics and genetics are also linked. Additionally, the recently created Teleost Alternative Splicing Database with alternative splicing data for catfish ESTs (17) is linked to cBARBEL.
cBARBEL is intended to serve as a central database for catfish researchers. In addition to the data components and tools described above, the database frontpage is updated with relevant links, news and meeting information of importance to the catfish research community.
Data availability
All of the data in cBARBEL is freely available. Users can contact the Auburn cBARBEL project team or email the corresponding author to request a specific subset of the data or to make suggestions about future database content and features.
FUTURE PLANS
cBARBEL is continuously updated to include newly generated data. These new data types are incorporated and linked to the existing data when appropriate. New data and features we anticipate adding to cBARBEL in the near-term include:
Catfish next-generation sequence data (454 and Illumina) utilizing GBrowse v2.0 from genome and transcriptome sequencing.
Additional annotation and batch sequence retrieval tools (BLAST, gene ontology annotation and primer design).
Expression data and links—microarray platform information and links to NCBI GEO-archived catfish data.
FUNDING
The USDA’s National Institute of Food and Agriculture; a scholarship from the Chinese Scholarship Council (to J.L.); and a scholarship from Alabama EPSCoR Graduate Research Scholar’s Program (to J.L.). Funding for open access charge: Discretionary Lab funds.
Conflict of interest statement. None declared.
ACKNOWLEDGEMENTS
We wish to thank members of the Catfish Genome Consortium for their efforts in generating catfish genome resources, especially members of the USDA-ARS Catfish Genetics Research Unit, Drs. Geoff Waldbieser and Sylvie Quiniou. We also thank NRSP-8′s Animal Genome Research Bioinformatics program (http://www.animalgenome.org/) for site hosting and assistance.
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Associated Data
This section collects any data citations, data availability statements, or supplementary materials included in this article.
Data Availability Statement
All of the data in cBARBEL is freely available. Users can contact the Auburn cBARBEL project team or email the corresponding author to request a specific subset of the data or to make suggestions about future database content and features.