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.
Project description:Experiment to estimate mutatational variance of gene expression in Drosophila melanogaster at two times in development using 12 mutation accumulation lines. Keywords = evolution Keywords = quantitative genetics Keywords = Drosophila Keywords = mutation Each of 12 lines measured 8 times in each of two stages. For each stage, the design has two hexagons with 6 interior connections (=hybridizations) (all except opposite nodes). Each line in one hexagon is connected to one line in the other hexagon and that line's opposite. Each line is also connected to itself twice in the other stage. Dyes are balanced.
Project description:The aims of this study were to determine the impact of different levels of RNA degradation as well as to ascertain if the gene expression profiles obtained from bladder washing correlates to that obtained from the related bladder tumor. We obtained tissue tumor and bladder washing tumor samples from the same individual. We degraded each intact RNA to 3 different degradation states with dye swap (2x4x2 arrays). All tumor RNAs were compared against a pool of 4 healthy control samples with intact RNA (named as C0).
Project description:http://www.nature.com/ng/journal/v33/n2/abs/ng1086.html. Little is known about broad patterns of variation and evolution of gene expression during any developmental process. Here we investigate variation in genome-wide gene expression among Drosophila simulans, Drosophila yakuba and four strains of Drosophila melanogaster during a major developmental transition?the start of metamorphosis. Differences in gene activity between these lineages follow a phylogenetic pattern, and 27% of all of the genes in these genomes differ in their developmental gene expression between at least two strains or species. We identify, on a gene-by-gene basis, the evolutionary forces that shape this variation and show that, both within the transcriptional network that controls metamorphosis and across the whole genome, the expression changes of transcription factor genes are relatively stable, whereas those of their downstream targets are more likely to have evolved. Our results demonstrate extensive evolution of developmental gene expression among closely related species. Extended from the original publication by updating annotation to release 4.0 and using 8 arrays for each strain/species. All Drosophila yakuba arrays are new. see also: PMID: 15122255. http://www.nature.com/ng/journal/v36/n6/abs/ng1355.html Gu Z, Rifkin SA, White KP, Li WH Duplicate genes increase gene expression diversity within and between species.
Project description:Aims of study: (1) To identify systemic differences in osteoarthritic (OA) bone that contribute to OA pathogenesis. (2) Identify novel osteoporotic (OP) bone-related disease genes. Study involved comparison of trabecular bone extracted from the intertrochanteric (IT) region of the proximal femur (PF) from OA, OP and normal/control (CTL) individuals. Bone was obtained from OA and OP individuals at surgery for total hip replacement and from CTL individuals at autopsy. Keywords: Bone tissue comparison, diseased versus non-diseased, factorial design, linear modelling Four sets of sample comparisons (39 comparisons in total) were made in this study. These comprised 10 OA-CTL female, 10 OA-CTL male, 10 OA-OP female and 9 OP-CTL female comparisons. A Compugen Human 19K-oligo library spotted onto glass slides by the Adelaide Microarray facility (AMF) was used in this study. The slides were interrogated by competitive hybridisation of Cy3 and Cy5 labelled pairs of OA-CTL, OA-OP or OP-CTL amplified RNA samples. Sample pairs were age-matched as closely as possible. A biological dye-swap strategy was employed. After hybridisation and washing of the slides they were scanned using a GenePix 4000B Scanner driven by GenePix Pro 4.0. All analyses were performed using the statistical programming and graphics environment R. The “SPOT” software package was used to identify spots by adaptive segmentation method and subtract backgrounds utilising morphological opening approach. Data analysis was performed in R using Bioconductor. Loess print tip method was used to correct for dye-bias and intensity within each group of adjacent spots printed by one pin. Linear modelling was performed with the Limma package of Bioconductor. Linear models were used to incorporate all available data into a single analysis. This allowed the use of both direct and indirect comparisons in calculation of expression ratios as well as improved accuracy in estimation of variance for each gene. Factorial design utilised the following factors: medical condition (OA or OP or CTL); sex (male or female) for OA patients only.
Project description:This is a time course experiment with 4 - 6 slides per time point where all samples were hybridised to time zero. The microarray slides were scanned using a GenePix 4000B Scanner (Axon Instruments, Foster City, CA, USA). The â??SPOTâ?? software package (CSIRO) was used to identify spots and subtract backgrounds. The extracted information was normalised using the â??Statistics in Microarray Analysis" (SMA) open source R package. Scaled print tip group lowes normalisation was performed for each slide and replicate slides were scaled to each other. The cDNAs used in this microarray were identified as part of collaborative project between the IMVS and Bionomics Limited. The project aims were to identify genes up-regulated during in vitro capillary tube formation as targets for angiogenesis-based therapeutics. This SuperSeries is composed of the following subset Series: GSE767: time 0.5 hr to 0 hr GSE771: time 3 hr to 0 hr GSE777: time 6 hr to 0 hr GSE778: time 24 hr to 0 hr Any requests for further information regarding the data generated from this collaboration should be directed to Bionomics Limited (www.bionomics.com.au) at the following address: Bionomics Limited 31 Dalgleish Street Thebarton, South Australia Australia, 5031 Phone: 618 8354 6104 Fax: 618 8354 6199 Email: busdev@bionomics.com.au Refer to individual Series
Project description:To understand better the mechanisms controlling the balance between proliferation and and differentiation during myelopoiesis we have utilized teh bi-potent FDB1 myeloid cell line which differentiates to granulocytes and macrophages in response to GM-CSF and thus provides a dissectable model to analyse the switch between growth and differentiation. This was further studied using this cell line in which a second site mutation Y577F had been generated. A factorial time-course design between the two cell lines and over time, combined with linear modelling and geneset enrichment analysis identified the expression changes associated with the switch between granulocyte differentiation and macrophage differentiation. We observed that a single intracellular tyrosine residue (Tyr577) mediates the granulocyte fate decision. The loss of granulocyte differentiation and enhanced macrophage differentiation observed in this second site mutant is associated with accumulation of Beta-Catenin and activation of Tcf4 and other Wnt Target genes. Factorial design. First Factor was cell type with levels FDB1 expressing FI-Delta receptor mutant and FDB1 expressing FI-Delta Y577F receptor mutant. Second factor is time with levels 0 and 72 hours. Two biological replicates wre prepared for each cell line and comparions were perfomed for each cell line and directly between cell lines at 0 and 72hs. Additional dye swaps were done with FI Delta and FIDelta Y577F at 0h and the FI Delta 0-72h comparison.
Project description:We used microarray analysis to screen the Drosophila genome for effects of Wolbachia infection on transcript levels in wildtype adult immature ovaries. Keywords: Comparative expression analysis of infection effect To generate the ovaries to be compared, we first rid a wildtype Canton-S line of Drosophila of the well characterized YW strain of Wolbachia that it harbored (Bourtzis et al., 1998, Presgraves, 2000) by raising the flies for three generations on food containing tetracycline. After the line had been raised for three more generations on food without tetracycline, we used a standard PCR assay (O'Neil et al., 1999) to confirm that the line was Wolbachia free. Genetically matched infected and uninfected females were recovered as the daughters from two reciprocal crosses between the infected and the cured Canton-S lines. Previtellogenic ovaries were harvested from these females 1-5 h after eclosion
Project description:We used chromatin immunoprecipitation (ChIP) in combination with human promoter microarrays to identify 216 putative SRF binding sites in the human genome. We performed independent quantitative PCR validation at over half of these sites that resulted in 146 sites we assert to be true binding sites at over 90% confidence. Nearly half of the sites are bound by SRF in only one of the three cell types we tested, providing strong evidence for the diverse roles for SRF in different cell types. Keywords: ChIP-chip SRF binding was tested in three different cell lines. It was validated by qPCR. Using the proximal promoter arrays, we are assaying binding in approximately 19,000 human promoters.