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A simplified platform to generate and purify human primordial germ cells in vitro provides new insights into germline specification

ABSTRACT: A simplified platform to generate and purify human primordial germ cells in vitro provides new insights into germline specification

PROVIDER: PRJNA867179 | ENA |

REPOSITORIES: ENA

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			Action	DRS
	SRR12588700.fastq.gz	Fastqsanger.gz
	SRR12588701.fastq.gz	Fastqsanger.gz
	SRR12588702.fastq.gz	Fastqsanger.gz
	SRR12588703.fastq.gz	Fastqsanger.gz
	SRR12588704.fastq.gz	Fastqsanger.gz

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Project description:Generating primordial germ cells (PGCs) from human pluripotent stem cells (hPSCs) advances studies of human reproduction and development of infertility treatments, but currently entails complex 3D aggregates. Here we develop a simplified, monolayer method to differentiate hPSCs into PGCs within 3.5 days, with higher efficiencies and improved consistency across multiple hPSC lines. We used our simplified differentiation platform, bulk RNA-seq and single-cell RNA-sequencing to uncover new insights into PGC specification. Transient WNT activation for 12 hours followed by WNT inhibition specified PGCs; by contrast, sustained WNT instead induced primitive streak. Thus, somatic (primitive streak) and PGCs are related—yet distinct—lineages segregated by temporally-dynamic signaling. Pluripotency factors are continuously expressed during the transition from pluripotency to posterior epiblast to PGCs, thus bridging pluripotent and germline states. Finally, hPSC-derived PGCs can be easily purified by virtue of their CXCR4+PDGFRA-GARP- surface-marker profile and single-cell RNA-sequencing reveals that they harbor strong transcriptional similarities with fetal PGCs. We report single-cell RNA-sequencing data of in vitro differentiated hPSCs (H9) cells into human Primordial germ cell-like cells (PGCLCs), profiled at different time-points of differentiation - Day 0 (H9), D0.5, D3.5 (sorted) and D3.5 unsorted population and a negative control of hPSCs differentiated into definitive endoderm (DE) (using the protocol described in (Loh et al., 2014, Cell Stem Cell 14, 237-252)). The data comprises of 5 single-cell RNA-sequencing libraries (H9, D0.5, D3.5S, D3.5U and DE) generated using 10xChromium Single cell RNA-expression platform and 10xGenomics Chromium version 2 chemistry with a targeted capture of ~6000 cells each. Single cell libraries were sequenced on Illumina HiSeq4000 platform as paired-end 150bp reads. Bulk RNA seq libraries were sequnced as paired-end reads.

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A simplified platform to generate and purify human primordial germ cells in vitro provides new insights into germline specification

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