Fish and chips: Various methodologies demonstrate utility of a 16,006-gene salmonid microarray

BMC Genomics, Sep 2005

Background We have developed and fabricated a salmonid microarray containing cDNAs representing 16,006 genes. The genes spotted on the array have been stringently selected from Atlantic salmon and rainbow trout expressed sequence tag (EST) databases. The EST databases presently contain over 300,000 sequences from over 175 salmonid cDNA libraries derived from a wide variety of tissues and different developmental stages. In order to evaluate the utility of the microarray, a number of hybridization techniques and screening methods have been developed and tested. Results We have analyzed and evaluated the utility of a microarray containing 16,006 (16K) salmonid cDNAs in a variety of potential experimental settings. We quantified the amount of transcriptome binding that occurred in cross-species, organ complexity and intraspecific variation hybridization studies. We also developed a methodology to rapidly identify and confirm the contents of a bacterial artificial chromosome (BAC) library containing Atlantic salmon genomic DNA. Conclusion We validate and demonstrate the usefulness of the 16K microarray over a wide range of teleosts, even for transcriptome targets from species distantly related to salmonids. We show the potential of the use of the microarray in a variety of experimental settings through hybridization studies that examine the binding of targets derived from different organs and tissues. Intraspecific variation in transcriptome expression is evaluated and discussed. Finally, BAC hybridizations are demonstrated as a rapid and accurate means to identify gene content.

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Fish and chips: Various methodologies demonstrate utility of a 16,006-gene salmonid microarray

Kristian R von Schalburg 2 Matthew L Rise 1 Glenn A Cooper 2 Gordon D Brown 2 A Ross Gibbs 2 Colleen C Nelson 0 William S Davidson 3 Ben F Koop 2 0 The Prostate Centre at Vancouver General Hospital , Gene Array Facility, Vancouver, British Columbia, V6H 3Z6 , Canada 1 Great Lakes WATER Institute, University of Wisconsin-Milwaukee , Milwaukee, WI, 53204 , USA 2 Centre for Biomedical Research, University of Victoria , Victoria, British Columbia, V8W 3N5 , Canada 3 Department of Molecular Biology and Biochemistry, Simon Fraser University , Burnaby, British Columbia, V5A 1S6 , Canada Background: We have developed and fabricated a salmonid microarray containing cDNAs representing 16,006 genes. The genes spotted on the array have been stringently selected from Atlantic salmon and rainbow trout expressed sequence tag (EST) databases. The EST databases presently contain over 300,000 sequences from over 175 salmonid cDNA libraries derived from a wide variety of tissues and different developmental stages. In order to evaluate the utility of the microarray, a number of hybridization techniques and screening methods have been developed and tested. Results: We have analyzed and evaluated the utility of a microarray containing 16,006 (16K) salmonid cDNAs in a variety of potential experimental settings. We quantified the amount of transcriptome binding that occurred in cross-species, organ complexity and intraspecific variation hybridization studies. We also developed a methodology to rapidly identify and confirm the contents of a bacterial artificial chromosome (BAC) library containing Atlantic salmon genomic DNA. Conclusion: We validate and demonstrate the usefulness of the 16K microarray over a wide range of teleosts, even for transcriptome targets from species distantly related to salmonids. We show the potential of the use of the microarray in a variety of experimental settings through hybridization studies that examine the binding of targets derived from different organs and tissues. Intraspecific variation in transcriptome expression is evaluated and discussed. Finally, BAC hybridizations are demonstrated as a rapid and accurate means to identify gene content. - Background Atlantic salmon are part of the Salmonidae family which comprise all salmon, trout, whitefish, grayling, and charr. A tremendous amount of basic biology is already known about salmonids from studies carried out on their physiology, population dynamics, behavioural ecology and phylogenetics [1]. Salmon also provide an excellent model system in which to study fundamental genetic mechanisms of growth, development, reproduction and response to infection and disease. For example, salmonids serve as prominent models for studies involving environmental toxicology [2], carcinogenesis [3], comparative immunology [4], the molecular genetics and physiology of the stress response [5], olfaction [6], vision [7], osmoregulation [8], growth [9] and gametogenesis [10]. Answers to fundamental scientific questions can also be gained from the study of salmonid genomes. The ancestor of all extant salmonids underwent a whole genome duplication and after a series of subsequent genetic events, salmon are now considered to be pseudo-tetraploid. How a genome reorganizes itself to cope with a duplicated genome and the importance of gene duplications for evolution and adaptation are long standing issues that remain unresolved. Questions regarding the origins of genomes have direct implication for our understanding of the roles of gene families, duplication and deletion of segments of genomes, and the mutational process in human health and disease. They also provide a foundation for understanding the genome of Atlantic salmon to benefit conservation and enhancement of wild stocks, aquaculture and environmental assessments. Genomic resources enable us to address fundamental scientific questions concerning the evolution of salmonid genomes, and the expression of genes and proteins in a wide variety of natural and altered environments and conditions. Toward these goals, more than 175 cDNA libraries have been constructed from a wide variety of tissues and different developmental stages and more than 300,000 salmonid cDNA sequence reads have been combined from a consortium comprising groups from Canada (Ben Koop et al. and the Genomics Research on Atlantic Salmon Project (GRASP); Susan Douglas et al. and the Institute for Marine Biosciences, NRC); France (Yann Guiguen et al. and INRA-SCRIBE); Norway (Bjorn Hoyheim et al. and the Norwegian School of Veterinary Science (NSVS)) and the U.S.A. (Caird Rexroad III and the USDA/ARS National Center for Cool and Cold Water Aquaculture). These sequences were assembled into over 40,000 unique contigs. A preliminary microarray of 3,557 cDNAs was constructed and assessed on its' ability to provide new data in the study of cellular and tissue responses to pollutants, diseases and stress, as well as for reproduction and development (...truncated)


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Kristian R von Schalburg, Matthew L Rise, Glenn A Cooper, Gordon D Brown, A Ross Gibbs, Colleen C Nelson, William S Davidson, Ben F Koop. Fish and chips: Various methodologies demonstrate utility of a 16,006-gene salmonid microarray, BMC Genomics, 2005, pp. 126, 6, DOI: 10.1186/1471-2164-6-126