Fischer Antje H. L.

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Fischer
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Antje H. L.
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  • Preprint
    SeaBase : a multispecies transcriptomic resource and platform for gene network inference
    ( 2014-05) Fischer, Antje H. L. ; Mozzherin, Dmitry ; Eren, A. Murat ; Lans, Kristen D. ; Wilson, Nathan ; Cosentino, Carlo ; Smith, Joel
    Marine and aquatic animals are extraordinarily useful as models for identifying mechanisms of development and evolution, regeneration, resistance to cancer, longevity and symbiosis, among many other areas of research. This is due to the great diversity of these organisms and their wide-ranging capabilities. Genomics tools are essential for taking advantage of these “free lessons” of nature. However, genomics and transcriptomics are challenging in emerging model systems. Here, we present SeaBase, a tool for helping to meet these needs. Specifically, SeaBase provides a platform for sharing and searching transcriptome data. More importantly, SeaBase will support a growing number of tools for inferring gene network mechanisms. The first dataset available on SeaBase is a developmental transcriptome profile of the sea anemone Nematostella vectensis (Anthozoa, Cnidaria). Additional datasets are currently being prepared and we are aiming to expand SeaBase to include user-supplied data for any number of marine and aquatic organisms, thereby supporting many potentially new models for gene network studies.
  • Dataset
    Nematostella vectensis BAC sequences
    ( 2013-07-11) Fischer, Antje H. L. ; Tulin, Sarah ; Fredman, David ; Smith, Joel
    A key tool for investigating the regulation of genes is represented by Bacterial Artificial Chromosomes-reporter constructs (BAC). BACs are large insert libraries, often >>100 kb, which thus capture the genomic sequences surrounding a gene of interest, including all, or nearly all, of the elements underpinning regulation. Recombinant BACs, containing a reporter gene in place of the endogenous coding sequence of genes, can be utilized to drive the expression of reporter genes under the regulatory control of the gene of interest while still embedded within its genomic context. Systematic deletions within the BAC reporter construct can be used to identify the minimal reporter in an unbiased way, avoiding the risk of overlooking regulatory elements that may be many kilobases away from the transcription start-site. Nematostella vectensis (Edwardsiidae, Anthozoa, Cnidaria) has become an important model in regenerative biology, ecology, and especially in studies of evo-devo and gene-regulatory networks due to its interesting phylogenetic position and amenability to molecular techniques. This dataset represents the sequencing of 5 Nematostella vectensis BACs containing the genes of interest: NvDpp, NvChordin, NvFoxa, NvWnta, and NvWnt1.
  • Dataset
    Nematostella High-density RNAseq time-course
    ( 2013-06-14) Fischer, Antje H. L. ; Smith, Joel
    RNA-Seq was performed on Nematostella embryos at 20 timepoints: unfertilized eggs, 1hour post fertilization (hpf), 2hpf, 3hpf, 4hpf, 5hpf, 6hpf, 7hpf, 8hpf, 9hpf, 10hpf, 11hpf, 12hpf, 13hpf, 14hpf, 15hpf, 16hpf, 17hpf, 18hpf, 19hpf. Two samples (A and B) were collected for each time point. All embryos were fertilized at the same time and 300 embryos were collected for each sample. After harvesting the embroys, they were lysed and the mRNAs was extracted using Dynabeads (Invitrogen). Directional sequencing libraries were constructed using the ScriptSeq V2 kit (Epicentre). The libraries were size selected with PippinPrep to 300bp insert size. Two different control RNA sets from the National Institute of Standards and Technology (NIST) (ERCC Spike-in, Ambion) were added into the m-RNA prior to the library preparation. Libraries were sequenced using the Illumina HiSeq 1000, version 3 chemistry, 100bp paired-end reads and produced over 1.4 billion reads. The reads were quality controlled filtered and mapped against the Nematostella embryonic transcriptome (Tulin et al.). This database contains the raw read files for each library.
  • Preprint
    Employing BAC-reporter constructs in the sea anemone Nematostella vectensis
    ( 2013-07-05) Fischer, Antje H. L. ; Tulin, Sarah ; Fredman, David ; Smith, Joel
    Changes in the expression and function of genes drive evolutionary change. Comparing how genes are regulated in different species is therefore becoming an important part of evo-devo studies. A key tool for investigating the regulation of genes is represented by Bacterial Artificial Chromosomes-reporter constructs (BAC). BACs are large insert libraries, often >>100 kb, which thus capture the genomic sequences surrounding a gene of interest, including all, or nearly all, of the elements underpinning regulation. Recombinant BACs, containing a reporter gene in place of the endogenous coding sequence of genes, can be utilized to drive the expression of reporter genes under the regulatory control of the gene of interest while still embedded within its genomic context. Systematic deletions within the BAC reporter construct can be used to identify the minimal reporter in an unbiased way, avoiding the risk of overlooking regulatory elements that may be many kilobases away from the transcription start-site. Nematostella vectensis (Edwardsiidae, Anthozoa, Cnidaria) has become an important model in regenerative biology, ecology, and especially in studies of evo-devo and gene-regulatory networks due to its interesting phylogenetic position and amenability to molecular techniques. The increasing interest in this rising model system also led to a demand for methods that can be used to study the regulation of genes in Nematostella. Here we present our progress in employing BAC reporter constructs to visualize gene-expression in Nematostella. Using a new Nematostella-specific recombination cassette, we made nine different BAC reporter constructs. Although five BAC recombinants gave variable effects, three constructs, namely Nv-bra:eGFP::L10 BAC, Nv-dpp:eGFP::L10 BAC, and Nv-grm:eGFP::L10 BAC, delivered promising results. We show that these three constructs express the reporter gene eGFP in 10.4% – 17.2% of all analyzed larvae, out of which 26.2 – 41.9% express GFP in a mosaic fashion within the expected domain. In addition to the expression within the known domains, we also observed cases of misexpression of eGFP and examples that could represent actual expression outside the described domain. Furthermore, we deep-sequenced and assembled five different BACs containing Nv-chordin, Nv-foxa, Nv-dpp, Nv-wnta, and Nvwnt1, to improve assembly around these genes. The use of BAC reporter constructs will foster cis-regulatory analyses in Nematostella and thus help to improve our understanding of the regulatory network in this cnidarian system. Ultimately, this will advance the comparison of gene-regulation across species and lead to a much better understanding of evolutionary changes and novelties.