Project description:This experiment assessed the natural gene expression variation present between colonies of the Indo-Pacific reef-building coral Acropora millepora, and additionally explored whether gene expression differed between two different intron haplotypes according to intron 4-500 in a carbonic anhydrase homolog. This study found no correspondence between host genotype and transcriptional state, but found significant intercolony variation, detecting 488 representing unique genes or 17% of the total genes analyzed. Such transcriptomic variation could be the basis upon which natural selection can act. Underlying variation could potentially allow reef corals to respond to different environments. Whether this source of variation and the genetic responses of corals and its symbionts will allow coral reefs to cope to the rapid pace of global change remains unknown. A. millepora colonies were brought to a common garden in the reef lagoon, i.e. under the same environmental conditions. This common garden combined with acclimatization removes environmental effects on the physiology of the coral colonies. For the comparison of the two intron haplotypes, we applied a multiple dye-swap microarray design for the two groups of coral colonies (N=3 per group) defined based on the two genotypes resolved with the use of intron 4-500 (Fig. 1). To also examine the intra-haplotype variation we added a loop design nested to the above multiple dye-swap design, where three samples per colony were included. Colonies 1, 2, and 3 are of intron 4-500 haplotype 1; colonies 4, 5, and 6 are haplotype 2.
Project description:Microarray technology provides a powerful tool for gene discovery studies, but the development of microarrays for individual species can be expensive and time-consuming. In this study, we test the suitability of a Danio rerio oligonucleotide microarray for application in a species with few genomic resources, the coral reef fish Pomacentrus moluccensis. Coral reef fishes are expected to experience rising sea surface temperatures due to climate change. How well tropical reef fish species will respond to these increased temperatures and which genes are important for resistance and adaptation to elevated temperatures is not known. Microarray technology may help identify candidate genes for thermal stress resistance in coral reef fishes. Results from a comparative genomic DNA hybridisation experiment and direct sequence comparisons indicate that for most genes there is significant sequence similarity between P. moluccensis and D. rerio, suggesting that the D. rerio array is applicable to P. moluccensis. Heterologous microarray experiments on heat-stressed P. moluccensis identified changes in transcript abundance at 120 gene loci, with many genes involved in protein processing, transcription, and cell growth. Changes in transcript abundance for a selection of candidate genes were confirmed by quantitative real-time PCR. We have demonstrated that heterologous microarrays can be successfully employed to study non-model organisms. Such a strategy thus greatly enhances the applicability of microarray technology to the field of environmental and functional genomics and will be useful for investigating the molecular basis of thermal adaptation in coral reef fishes. Keywords: stress response, comparative genomic hybridization (CGH) Common reference design [Stress response_P. moluccensis]: four individual treatment fish (heat-stressed) are contrasted in four microarray hybridisations against a pooled control consisting of four fish kept at ambient temperature. All eight fish employed in this analysis were wild-captured and are biological replicates. The experiment included dye-swap, i.e. stressed fish were labelled red in two hybridisations and green in the other two hybridisations. Common reference design [CGH_P. moluccensis and D. rerio]: four individual P. moluccensis gDNA samples are contrasted in four microarray hybridisations against a pooled gDNA sample consisting of three D. rerio. The experiment included dye-swaps.