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Accession IconSRP093264

Rapid molecular profiling of defined cell types using viral TRAP

Organism Icon Mus musculus
Sample Icon 28 Downloadable Samples
Technology Badge IconIllumina HiSeq 2500

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Description
Translational profiling methodologies enable the systematic characterization of cell types in complex tissues such as the mammalian brain, where neuronal isolation is exceptionally difficult. Here, we report a versatile strategy to profile CNS cell types in a spatiotemporally-restricted fashion by engineering a Cre-dependent adeno-associated virus expressing an EGFP-tagged ribosomal protein (AAV-FLEX-EGFPL10a) to access translating mRNAs by TRAP. We demonstrate the utility of this AAV to target a variety of genetically and anatomically defined neural populations expressing Cre recombinase and illustrate the ability of this viral TRAP (vTRAP) approach to recapitulate the molecular profiles obtained by bacTRAP in corticothalamic neurons across multiple serotypes. Furthermore, spatially restricting AAV injections enabled the elucidation of regional differences in gene expression within this cell type. Taken together, these results establish the broad applicability of the vTRAP strategy for the molecular dissection of any CNS or peripheral cell type that can be engineered to express Cre. Overall design: Polysome-bound mRNAs from TRAP IPs were compared to whole tissue mRNAs. Data was collected from MCH neurons in hypothalamus using vTRAP, cortical layer 6 Ntsr1 neurons using vTRAP, and cortical layer 6 Ntsr1 neurons using bacTRAP. We include vTRAP data from three AAV serotypes for the cortical Ntsr1 cells. We collected three replicates for IP and inputs for vTRAP experiments, while bacTRAP data was collected in duplicate.
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28
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