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Single-cell RNA-seq of sorted E11.5 AGM HSCs

ABSTRACT: Single-cell RNA-seq of sorted E11.5 AGM HSCs

PROVIDER: PRJNA578153 | ENA |

REPOSITORIES: ENA

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Single-cell RNA-seq of sorted E11.5 AGM HSCs

Project description:The aim of this study was to analyse the heterogeneity within the first haematopoietic stem cells generated during mouse development using single-cell RNA-Seq. Freshly dissected aorta-gonads-mesohenphros (AGM) regions from E11.5 mouse embryos were dissociated by collagenase and the cells stained for the cell surface markers EPCR and CD45. EPCR+CD45+ cells were index-sorted as single cells into 96-well plates and processed based on the SmartSeq2 protocol. PCA and tSNE analyses identified 2 main subpopulations which are segregated based on differences in cell cycle. Known HSC markers such as Gata2 and Runx1 show little correlation.

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Single-cell transcriptomic interrogation of hematopoietic progenitors within the E11.5 AGM of the mouse

Project description:To clarify the relationship between eMPPs and other progenitors, including pre-HSCs, within the AGM landscape, we performed single cell RNA-sequencing of hematopoietic progenitors within the E11.5 AGM.

2022-02-10 | GSE180357 | GEO

Transcriptomic analysis of E11.5 AGM VE cadherin populations

Project description:We sorted endothelial (EC), hematopoietic (HC) and hematopoietic stem/progenitor (HSPC) cell populations from AGM of E11.5 C57Bl6 embryos, established and compared their transcriptome to highlight specific regulators of hematopoietic emergence.

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ChIP-on-chip with anti-RBPj antibody from embryonic AGM E11.5

Project description:Hematopoietic Stem Cells (HSC) are originated during embryonic development from endothelial-like cells located in the ventral side of the dorsal aorta around day E10-12 of murine development. This region is called AGM for Aorta/Gonad/Mesonephros refering to the tissues around the hemogenic aorta. Hematopoiesis depends on the Notch pathway and the identification of Notch-targets is important for the understanding of blood origin. Hematopoietic Stem Cells (HSCs) specification occurs in the embryonic aorta and requires Notch activation, however which are the elements regulated by Notch that control this process are mainly unknown. Here, we took a genome-wide approach to identify putative direct Notch targets by precipitating the chromatin that binds to the Notch partner RBPj in the Aorta-Gonad-Mesonephros (AGM) tissue from E11.5 mouse embryos. This assay revealed 701 gene promoter regions as candidates to be regulated by Notch in the AGM. Chromatin was obtained from a pool of 40 dissected AGMs at E11.5. Chromatin immunoprecipitation (ChIP) was performed as previously described (Aguilera et al, PNAS 2004) with minor modifications. In brief, cross-linked chromatin was sonicated for 10 minutes, medium-power, 0.5-interval; with a Bioruptor (Diagenode) and precipitated with anti-RBPJ (Chu and Bresnick, 2004). After crosslinkage reversal, DNA was used as a template for PCR or for array hybridization. Mouse promoter chip on chip microarray SET (Agilent) was used to identify RBPj targets. It covers 70,000 best identified gene regions with a-5.5 kb to + 2.5 kb range, and has on average 25 probes per gene with an average probe to probe distance of 200 bp. The ChIP-on-chip was performed with dye swaps and one IgG control was brought along. Enrichment analysis was done by comparing the precipitation normalized dye swap signal with input control signal.

2014-10-22 | E-GEOD-52094 | biostudies-arrayexpress

Transcriptomic analysis of E11.5 AGM VE cadherin populations

Project description:Transcriptomic analysis of E11.5 AGM VE cadherin populations

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Project description:RNA-Seq of sorted E10.5 and E11.5 AGM cell populations from Gata3-GFP embryos

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Single-cell transcriptomic interrogation of hematopoietic progenitors within the E11.5 AGM of the mouse

Project description:Single-cell transcriptomic interrogation of hematopoietic progenitors within the E11.5 AGM of the mouse

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RNA-Seq of sorted E10.5 and E11.5 AGM cell populations from Gata3-GFP embryos

Project description:The aim of this study was to examine the whole transcriptome of pools of 20 cells of Gata3-expressing AGM subpopulations (haematopoietic [HC], mesenchymal [MC], sympathetic nervous system [SNS], and endothelial [EC]) and compare them to their non-Gata3 expressing counterparts

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AGM hematopoietic stem cells are differentially regulated by BMP and Hedgehog signalling pathways

Project description:The first HSCs are produced in the aorta-gonadmesonephros (AGM) region of the embryo through endothelial to a hematopoietic transition. BMP4 and Hedgehog affect their production/expansion, but it is unknown whether they act to affect the same HSCs. In this study using the BRE GFP reporter mouse strain that identifies BMP/Smad-activated cells, we find that the AGM harbors two types of adult-repopulating HSCs upon explant culture. Embryonic day 11 AGM are dissected and either analyzed directly, or after explant culture in conditions containing BMP/Hedgehog with or without cyclopamine. EC: endothelial enriched (CD31+Kit-); MC: mesenchymal cell enriched (CD31-Kit-); HPSC: hematopoietic progenitor/stem cell enriched; AGM11: E11 fresh AGMs; AGMex: AGM after explant culture; AGMcy: AGM after explant in presence of cyclopamine; CD31p: CD31 positive; CD31n: CD31 negative; KITp: c-Kit positive; KITn: c-Kit negative; BREp: BRE-GFP positive; BREn: BRE-GFP negative

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Transcriptome imposed by Notch ligands (Jag1 or Dll4) on ckit+ and ckit- cells from E11.5 AGM

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