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Transcriptome profiling of purified red blood cells from Nxf1 mutant and control mice [Erythrocytes]

ABSTRACT: Transcriptome profiling of purified red blood cells from Nxf1 mutant and control mice [Erythrocytes]

PROVIDER: PRJNA592453 | ENA |

REPOSITORIES: ENA

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			Action	DRS
	SRR10559878_1.fastq.gz	Fastqsanger.gz
	SRR10559878_2.fastq.gz	Fastqsanger.gz
	SRR10559879_1.fastq.gz	Fastqsanger.gz
	SRR10559879_2.fastq.gz	Fastqsanger.gz
	SRR10559880_1.fastq.gz	Fastqsanger.gz

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Transcriptome profiling of purified red blood cells from Nxf1 mutant and control mice [Erythrocytes]

Project description:Purpose: Nxf1 is thought to be an essential nuclear exporter of messenger RNA (mRNA) in eukaryotic cells. Whether perturbations in the Nxf1 pathway affect mammalian physiology is not known. The aim of this study is to determine the impact of a Nxf1 mutation in the representation of mRNA transcripts in erythrocytes. Methods: Red blood cells were obtained from cardiac puncture, mixed with 100µl Aster Jandl citrate-based anticoagulant and subsequently diluted in 2 ml of phosphate buffer saline (PBS) with 2mM EDTA. This suspension was layered with 1.5 ml Ficoll-paque PLUS (GE Healthcare) and centrifuged at 400g for 30 minutes at room temperature with minimal acceleration and brake. The red blood cell containing pellet was washed twice with PBS 2mM EDTA and lysed for RNA extraction. The purity of each sample was checked by flow cytometry to assess the relative contamination of Ter199+ red blood cells with CD41+ platelets, CD45+ lymphocytes and DRAQ5+ (Thermo Fisher Scientific) nucleated cells. Red blood cell suspensions used for RNAseq were > 99.8% pure. RNA was extracted with Qiagen RNA extraction kit.

2020-03-04 | GSE141160 | GEO

Transcriptome profiling of purified mouse platelets from Nxf1 mutant and control mice [Platelet]

Project description:Purpose: Nxf1 is thought to be an essential nuclear exporter of messenger RNA (mRNA) in eukaryotic cells. Whether perturbations in the Nxf1 pathway affect mammalian physiology is not known. The aim of this study is to determine the impact of a Nxf1 mutation in the representation of mRNA transcripts in platelets. Methods: Platelets were purified from individual males. Blood was obtained by cardiac puncture into 0.1 volume of Aster Jandl citrate-based anticoagulant. Mouse platelet rich fraction was obtained by centrifugation of the murine blood at 125 g for 8 min at room temperature, followed by centrifugation of the supernatant buffy coat at 125 g for 8 min. Mouse platelets were washed by two sequential centrifugations at 860 g for 5 min in 140 mM NaCl, 5 mM KCl, 12 mM trisodium citrate, 10 mM glucose, and 12.5 mM sucrose, pH 6.0.The platelet pellet was resuspended in 10 mM Hepes, 140 Mm NaCl, 3 mM KCl, 0.5 mM MgCl2, 10 mM glucose, and 0.5 mM NaHCO3, pH 7.4. The purity of each platelet suspension was assessed by flow cytometry and suspensions for which more than 98% of total events were CD41+ platelets were pooled together. RNA was purified with Norgen cytoplasmic and nuclear fractionation RNA purification kit.

2020-03-04 | GSE141159 | GEO

Transcriptome profiling of purified mouse platelets from Nxf1 mutant and control mice [Platelet]

Project description:Transcriptome profiling of purified mouse platelets from Nxf1 mutant and control mice [Platelet]

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Transcriptome profiling of cytoplasmic fraction of naïve CD8 T cells from Nxf1 mutant and control mice [Lymphocytes]

Project description:Purpose: Nxf1 is thought to be an essential nuclear exporter of messenger RNA (mRNA) in eukaryotic cells. Whether perturbations in the Nxf1 pathway affect mammalian physiology is not known. The aim of this study is to determine the impact of a Nxf1 mutation in the representation of mRNA transcripts in the cytoplasm of naïve CD8 T cells. Methods: For Naïve CD8 T cell isolation, splenocyte suspensions were submitted to red blood cell lysis, stained first with biotinylated CD8 antibodies and then labelled with anti-Biotin microbeads. CD8+ T cells were magnetically enriched by positive selection on MACS separation columns, stained with fluorescently-labelled antibodies before being FACS-sorted as CD8+ CD44low CD62L+. Sorted Naïve T cells were fractionated with Norgen cytoplasmic and nuclear fractionation RNA purification kit. Only cytoplasmic fractions were used for subsequent RNAseq.

2020-03-04 | GSE141158 | GEO

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Project description:Erythrocytes are the most numerous cells of the human body (Sender 2016). Their lifespan in the blood stream is around 120 days. The turnover implies the production of 2 million new reticulocytes each second. Circulating reticulocytes achieve their maturation in 2 to 3 days. During this short time, important modifications occur such as membrane remodeling, removing of remaining organelles and specific elimination of useless proteins such as the transferrin receptor. This remodeling will ensure the optimization of the erythrocyte ability to fulfil its transport functions. We performed a deep label-free quantitative proteome analysis of purified erythrocytes and reticulocytes to better understand the physiology of the erythrocytes and the last steps of their maturation.

2018-09-18 | PXD009258 | Pride

Transcriptome profiling of blood cells from Nxf1 mutant and control mice

Project description:Transcriptome profiling of blood cells from Nxf1 mutant and control mice

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