Several studies have highlighted the changes in the gene expression due to the hypoxia response in fishes, but the systematic organization of the information and the analytical platform for such genes are lacking. In the present study, an attempt was made to develop a database of hypoxia responsive genes in fishes (HRGFish), integrated with analytical tools, using LAMPP technology. Genes reported in hypoxia response for fishes were compiled through literature survey and the database presently covers 818 gene sequences and 35 gene types from 38 fishes. The upstream fragments (3,000 bp), covered in this database, enables to compute CG dinucleotides frequencies, motif finding of the hypoxia response element, identification of CpG island and mapping with the reference promoter of zebrafish. The database also includes functional annotation of genes and provides tools for analyzing sequences and designing primers for selected gene fragments. This may be the first database on the hypoxia response genes in fishes that provides a workbench to the scientific community involved in studying the evolution and ecological adaptation of the fish species in relation to hypoxia.

BackgroundMicrosatellite DNA is one of many powerful genetic markers used for the construction of genetic linkage maps and the study of population genetics. The biological databases in public domain hold vast numbers of microsatellite sequences for many organisms including fishes. The microsatellite data available in these data sources were extracted and managed into a database that facilitates sequences analysis and browsing relevant information. The system also helps to design primer sequences for flanking regions of repeat loci for PCR identification of polymorphism within populations.DescriptionFishMicrosat is a database of microsatellite sequences of fishes and shellfishes that includes important aquaculture species such as Lates calcarifer, Ctenopharyngodon idella, Hypophthalmichthys molitrix, Penaeus monodon, Labeo rohita, Oreochromis niloticus, Fenneropenaeus indicus and Macrobrachium rosenbergii. The database contains 4398 microsatellite sequences of 41 species belonging to 15 families from the Indian subcontinent. GenBank of NCBI was used as a prime data source for developing the database. The database presents information about simple and compound microsatellites, their clusters and locus orientation within sequences. The database has been integrated with different tools in a web interface such as primer designing, locus finding, mapping repeats, detecting similarities among sequences across species, and searching using motifs and keywords. In addition, the database has the ability to browse information on the top 10 families and the top 10 species, through record overview.ConclusionsFishMicrosat database is a useful resource for fish and shellfish microsatellite analyses and locus identification across species, which has important applications in population genetics, evolutionary studies and genetic relatedness among species. The database can be expanded further to include the microsatellite data of fishes and shellfishes from other regions and available information on genome sequencing project of species of aquaculture importance.

Cyprinus carpio is world’s most widely distributed freshwater species highly used in aquaculture. It is a hypoxia-tolerant species as it lives in oxygen-deficient environment for a long period. The tolerance potential of an animal against hypoxia relates it to induced gene expression, where a hypoxia-inducible factor (HIF) binds to a transcriptionally active site, hypoxia response element (HRE), a 5-base short motif that lies within the promoter/enhancer region of a certain gene, for inducing gene expression and preventing/minimizing hypoxia effects. HRE is functionally active when it contains another motif, the hypoxia ancillary sequence (HAS), which is typically adjacent to downstream of HRE within 7- to 15-nt space. Here, an attempt was made for mining HRE and identifying functional HIF binding sites (HBS) in a genome-wide analysis of C. carpio . For this, gene information along with the 5,000-nt upstream (−4,900 to +100) sequences of 31,466 protein coding genes was downloaded from “Gene” and “RefSeq” databases. Analysis was performed after filtration of the impracticable genes. A total of 116,148 HRE consensus sequences were mined from 29,545 genes in different promoter regions. HRE with HAS consensus motifs were found in the promoter region of 9,589 genes. Further, the already reported genes for hypoxia response in humans and zebrafish were reanalyzed for detecting HRE sites in their promoters and used for comparative analysis with gene promoters of C. carpio for providing support to identify functional HBS in the gene promoter of C. carpio . An interactive user interface HREExplorer was developed for presenting the results on the World Wide Web and visualizing possible HBS in protein coding genes in C. carpio and displaying the comparative results along with the reported hypoxia-responsive genes of zebrafish and reported hypoxia-inducible genes in humans. In this study, a set of Perl program was written for the compilation and analysis of information that might be used for a similar study in other species. This novel work may provide a workbench for analyzing the promoter regions of hypoxia-responsive genes.

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