Project description:Transcriptomes of organisms reveal differentiation associated with the use of different habitats. However, this leaves open how much of the observed differentiation can be attributed to genetic differences or to transcriptional plasticity. In this study, we disentangle causes of differential gene expression in larvae of the European fire salamander from the Kottenforst forest in Germany. Larvae inhabit permanent streams and ephemeral ponds and represent an example of a young evolutionary split associated with contrasting ecological conditions. We found ample evidence for differentiation among larvae occupying different habitats in nature with 2800 out of 11797 genes being differentially expressed based on transcriptome data from salamander sampled in their natural habitat (see GEO Series GSE100819). We then quantified transcriptional plasticity towards temperature and genetic differentiation based on controlled temperature laboratory experiments. Gene-by-environment interactions modelling revealed that 28 % of the gene expression divergence observed among samples in nature could be attributed to plasticity related to water temperature. Expression patterns of only a small number of 101 genes were affected by the genotype. Our analysis demonstrates that effects of environmental factors must be taken into account to explain variation of gene expression in salamanders in nature. Notwithstanding, it provides first evidence that genetic factors determined gene expression divergence between pond and stream ecotypes and could be involved in adaptive evolution.
Project description:A custom 8x60 k expression microarray for larvae of European fire salamander (Salamandra salamandra) was designed based on transcriptome sequencing. It is known the fact, that oligonucleotide probes differ in the binding behavior towards their target sequences. Therefore, we performed a calibration of our microarray where we assessed the binding behavior of the individual probes empirically. This information was used to normalize gene expression data measurements with the same microarray in another experiment. Please refer to the accompanying publication (Czypionka et al. 2015." Ecological transcriptomics – a non-lethal sampling approach for endangered fire salamanders" Methods in Ecology and Evolution) for more information. Labeled cRNA was prepared from Salamander larvae kept at 9°C and 17°C. A cRNA calibration pool was prepared with equimolar amounts of cRNA prepared from (a) a larvae (temperature: 9°C: source: pond KOE), (b) a larvae (temperature: 17°C: source: pond KOE), (c) a larvae (temperature: 9°C: source: stream KoGB (Klufterbach) and (d) a larvae (temperature: 17°C: source: stream KoGB (Klufterbach). See Steinfartz et al. (2007) (doi: 10.1111/j.1365-294X.2007.03490.x) for information of the source populations. Increasing amounts of labeled cRNA (75 ng, 150 ng, 300 ng, 600 ng, 1000 ng, 1400 ng, 1800 ng, 2200 ng), corresponding to (1/8, 1/4, 1/2, 1, 1 2/3, 2 1/3, 3 and 3 3/3 times the recommended amount of 600 ng) were hybridized to 8 microarrays (one microarray per dilution). The change in observed signal intensity in relation to the change in amount of labeled cRNA was used to infer the target-binding behavior of the individual probes. This information was extracted, to be used for a normalization procedure in another experiment with the same microarray (see Czypionka et al. 2015." Ecological transcriptomics – a non-lethal sampling approach for endangered fire salamanders" Methods in Ecology and Evolution). The current study provides only raw data for a calibration experiment, to validate the binding behavior of the different probes on a newly designed microarray for a non model organism (European Fire salamander). This calibration is based only on raw data. More information on targeted genes is provided in a different GEO dataset (currently submitted), where biological meaningful analysis are performed with data which are normalized based on this calibration.
Project description:Tadpoles of the anuran species Rana pirica can undergo predator-specific morphological responses. Exposure to a predation threat by larvae of the salamander Hynobius retardatus results in formation of a bulgy body (bulgy morph) with a higher tail. The objective of the present study was to use Affymetrix Xenopus Genechip to profile gene expression in the tail tissue by different predation threat. Tadpoles of Rana pirica treated with larvae salamander for 8days (brainS1, brainS2, brainS3) were analyzed with triplicate. Controls were cultured for 8days without larvae salamander (brainC1,brainC2,brainC3,brainC4,brainC5,brainC6). Brains from tadpoles after 8days of each treatment were dissected for RNA extraction and gene expression analysis using Affymetrix Xenopus Genechip arrays.
Project description:Time-course transcriptional profiling of rice leaf of the temperature shift experiment in a growth chamber. This experiment was performed to validate the results of field transcriptomic modeling.
Project description:Plethodontid salamanders are the largest family of salamanders and are classic models for studying the effect of rapidly evolving courtship pheromones on mating behavior and reproductive success. Despite interests in plethodontid reproduction, very little is known about the molecular composition of salamander gametes, as the extraordinary sizes of their genomes have impaired the development of various omic-scale resources. To identify what proteins may be expressed in salamander sperm, we performed DIA-MS on sperm samples from two plethodontid species, Plethodon shermani and Desmognathus ocoee. As the first detailed study of salamander sperm, this study partially fills in a critical taxonomic gap in the study of fertilization proteins in vertebrates.
Project description:Microarrays have evolved from low-density cDNA or oligonucleotide arrays to high-density platforms, for several study species even covering the complete transcriptome. At the same time, transcriptomics experiments have become more complex and multifactorial in nature, requiring many microarrays to assess multiple biologically relevant hypotheses. Scientists using this technology are therefore painfully aware of the high financial cost of a typical microarray experiment. Unfortunately, this often leads to either a suboptimal experimental design in an effort to reduce the cost by using fewer microarrays, or to abandoning microarray technology altogether. In this study, we argue that for many studies high-density full genome microarrays are in fact technical overkill. By selectively reducing full genome probe sets to a lower number of probes, it is possible to significantly reduce the total cost of a microarray experiment. The study consists of four microarray analyses: a cadmium probe selection experiment, a temperature probe selection experiment, a cadmium validation experiment and a cadmium validation experiment.
Project description:Tadpoles of the anuran species Rana pirica can undergo predator-specific morphological responses. Exposure to a predation threat by larvae of the salamander Hynobius retardatus results in formation of a bulgy body (bulgy morph) with a higher tail. Whereas, dragon fly also induced higher tail tadpole. The tadpoles revert to a normal phenotype upon removal of the larval salamander or dragon fly threat. The objective of the present study was to use Affymetrix Xenopus Genechip to profile gene expression in the tail tissue by different predation threat.