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Single-cell RNA-sequencing of Platynereis dumerilii

ABSTRACT: Single-cell RNA-sequencing of Platynereis dumerilii

PROVIDER: PRJEB22558 | ENA |

REPOSITORIES: ENA

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			Action	DRS
	ERR2115660_1.fastq.gz	Fastqsanger.gz
	ERR2115660_2.fastq.gz	Fastqsanger.gz
	ERR2115670_1.fastq.gz	Fastqsanger.gz
	ERR2115670_2.fastq.gz	Fastqsanger.gz
	ERR2115680_1.fastq.gz	Fastqsanger.gz

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Single-cell RNA-sequencing of Platynereis dumerilii

Project description:For unbiased, whole-organism wide cell type profiling, we randomly sampled cells from dissociated Platynereis larvae. To generate the single-cell mRNA-sequencing data, P. dumerilii larvae were dissociated, followed by cell capture, cDNA synthesis and amplification on the C1 Single-Cell Auto Prep IFCs for 5-10 um or 10-17 um cells (Fluidigm). Sequencing libraries were produced using Nexera XT DNA kit (Illumina). In total, we sequenced 596 samples, of which 373 correspond to single, alive cells that passed the quality check criteria. Part of this dataset was previously published (ArrayExpress accession number E-MTAB-2865). Here, we publish additional 383 sequenced cells.

2017-09-18 | E-MTAB-5953 | biostudies-arrayexpress

Single-cell RNA-sequencing of Platynereis dumerilii larval brain cells

Project description:Understanding cell type identity in complex tissues or organisms requires integration of each cell's expression profile with its spatial location within the tissue under study. We developed a high-throughput method that combines in vitro single-cell RNA-sequencing with a gene expression atlas to map single cells back to their location within the tissue of interest. We used the developing brain of a marine annelid, Platynereis dumerilii that is an important model system for studying bilaterian brain evolution, to benchmark our approach. To generate the single-cell mRNA-sequencing data, P. dumerilii larval brains were dissociated, followed by cell capture, cDNA synthesis and amplification on the C1 Single-Cell Auto Prep IFC for 10-17 um cells (Fluidigm). Sequencing libraries were produced using Nexera XT DNA kit (Illumina). In total, we sequenced 213 samples, of which 129 correspond to single, alive cells (as judged by visual inspection of the captured cells) with the remainder consisting of a variety of single dead cells (n=18), wells containing extracellular matrix contaminants (n=8) or multiple cells (n=17), as well as a negative controls where no cells were observed (n=41). For this dataset, we achieved ~90% success rate for the spatial mapping of the single-cell RNA-seq data to P. dumerilii brain atlas. NOTE: 72 additional samples were added on 13th December 2014.

2014-12-13 | E-MTAB-2865 | biostudies-arrayexpress

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Project description:The marine annelid Platynereis dumerilii has become a model system for evo-devo, neurobiology and marine biology. The functional assessment of its cell types, however, has so far been very limited. Here we report on the establishment of a generally applicable, cell type specific ablation technique to overcome this restriction. Using a transgenic strain expressing the bacterial enzyme nitroreductase (ntr) under the control of the worm's r-opsin1 locus, we show that the demarcated photoreceptor cells can be specifically ablated by the addition of the prodrug metronidazole (mtz). TUNEL staining indicates that ntr expressing cells undergo apoptotic cell death. As we used a transgenic strain co-expressing ntr with enhanced green fluorescent protein (egfp) coding sequence, we were able to validate the ablation of photoreceptors not only in fixed tissue, using r-opsin1 riboprobes, but also by monitoring eGFP+ cells in live animals. The specificity of the ablation was demonstrated by the normal presence of the eye pigment cells, as well as of neuronal markers expressed in other cells of the brain, such as phc2, tyrosine hydroxylase and brn1/2/4. Additional analyses of the position of DAPI stained nuclei, the brain's overall neuronal scaffold, as well as the positions and projections of serotonergic neurons further confirmed that mtz treatment did not induce general abnormalities in the worm's brain. As the prodrug is administered by adding it to the water, targeted ablation of specific cell types can be achieved throughout the life of the animal. We show that ablation conditions need to be adjusted to the size of the worms, likely due to differences in the penetration of the prodrug, and establish ablation conditions for worms containing 10 to 55 segments. Our results establish mtz/ntr mediated conditional cell ablation as a powerful functional tool in Platynereis.

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