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Expression profiling of FACS-purified fetal germ cells

ABSTRACT: We used microarrays to profile global gene expression changes of Pou5f1-GFP-positive germ cells between E11.5 to E15.5.

ORGANISM(S): Mus musculus

PROVIDER: GSE38126 | GEO | 2013/01/25

SECONDARY ACCESSION(S): PRJNA167379

REPOSITORIES: GEO

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Expression profiling of FACS-purified fetal germ cells

Project description:We used microarrays to profile global gene expression changes of Pou5f1-GFP-positive germ cells between E11.5 to E15.5. Germ cells were FACS-purified from gonadal single cell suspension based on Pou5f1-GFP expression. Three timepoints were included in this study: E11.5 (male/female), E13.5 (male) and E15.5 (male). For each timepoint, three biological replicates were analyzed. The Pou5f1-GFP-negative (non-germ cell) fraction of E13.5 (male) gonads was also included as a control.

2013-01-25 | E-GEOD-38126 | biostudies-arrayexpress

In vivo epigenomic profiling of germ cells reveals germ cell molecular signatures [ChIP-Seq]

Project description:The limited number of in vivo germ cells poses an impediment to genome-wide studies. Here, we applied a small-scale ChIP-Seq method on purified mouse fetal germ cells to generate genome-wide maps of four histone modifications (H3K4me3, H3K27me3, H3K27ac and H2BK20ac), facilitating the identification of active and repressed cis-regulatory elements in germ cells in vivo. Comparison of active chromatin state between somatic, embryonic stem cells (ESC) and germ cells revealed promoters and enhancers needed for stem cell maintenance and germ cell development. The nuclear receptor Nr5a2 motif is enriched at a subset of cis-regulatory regions and we confirm its role in germ cell differentiation. Interestingly, germ cells have comparatively more H3K27me3-marked sites that are absent in ESC and other somatic cell types. These repressed regions are enriched for retrotransposons and MHC genes and this indicates that these loci are specifically silenced in germ cells. Together, our study provides the first genome-wide histone modification maps of in vivo germ cells and revealed the molecular chromatin signatures unique to germ cells. Germ cells were FACS-purified from gonadal single cell suspension based on Pou5f1-GFP expression. ChIP-seq of Histone modification was done for two timepoints in this study: E11.5 (male/female), E13.5 (male). For E13.5 timepoint, two biological replicates were analyzed. In order to validate small scale ChIP-seq method limited number of ES cells were used to check consistency of ChIP-seq data.

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Effect of c-Maf mutation on Mafb-GFP-positive FACS-purified cells in the E12.5 mouse gonad

Project description:Morphogenesis of the gonad requires cell-cell adhesion changes between diverse cell types. In the Drosophila gonad, the gene traffic jam regulates cell adhesion changes required for gonad formation and germ cell development (Li et al., 2003. Nature Cell Biol). To determine if the mammalian homologs of traffic jam in mammals, c-Maf and Mafb, also play a role in the transcription regulation of cell adhesion molecules in the mouse gonad, we performed a microarray analysis of FACS-purified Mafb-GFP-positive cells in E12.5 male control and c-Maf/Mafb mutant gonads. We used microarrays to determine genes affected by c-Maf mutation in E12.5 mouse gonad/mesonephros interstitial cells and macrophages E12.5 XY control (c-Maf+/-;Mafb-GFP+/-) and c-mutant (c-Maf-/-;Mafb-GFP+/-) gonad/interstitial interstitial cells and macrophages were obtained by FACS sorting of Mafb-GFP-positive cells. RNA was extracted for subsequent hybridization on Affymetrix microarrays.

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Effect of c-Maf mutation on Mafb-GFP-positive FACS-purified cells in the E12.5 mouse gonad

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ATAC-seq of zebrafish primordial germ cells and somatic cells during early embryogenesis

Project description:Zebrafish Primordial Germ Cells (PGCs) were tracked in the Tg(Buc-GFP) line where the germ plasm protein Buc is fused to GFP. GFP-positive cells were isolated via FACS and RNA-seq was performed on polyadenylated transcripts for PGCs and somatic cells at 256-cell, high, dome, 10 somites and prim-5 stages. Two hundred cells were used for each biological replicate.

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Project description:To find the genes with H3K36me3 modificaiton at different stages of pancreas development, we isolated the pancreatic epithelial cells from E11.5, E13.5 and E15.5 fetal pancreas. Considering the low expression of H3K36me3 at E11.5, E11.5 samples worked as a "negantive" control. By compared with the peaks of E11.5, we was able to find genes under direct regulation of H3K36me3 at E13.5 and E15.5.

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