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| Status |
Public on Sep 22, 2021 |
| Title |
Scalable Production of Mouse Erythroid Cells using an in vitro Embryoid Body differentiation System [Capture-C] |
| Organism |
Mus musculus |
| Experiment type |
Other
|
| Summary |
Mouse embryonic stem (mES) cells can be manipulated ex-vivo to recapitulate the process of erythropoiesis, whereby multipotential haematopoietic stem cells undergo lineage specification, differentiation and maturation to produce functional erythroid cells. Although very useful for identifying specific progenitors and precursors, this cell system has not been fully exploited as a source of cells to analyse erythropoiesis, compared with primary erythroid cells or immortalised cell lines. Here we have established a protocol in which erythroid cells can be isolated in a scalable manner from differentiated embryoid bodies (EBs). Using transcriptional and epigenetic analysis, we show that all purified erythroid cells in this system are derived from the primitive haematopoietic lineage. Finally, we show that this system faithfully recapitulates normal erythropoiesis in the embryonic mouse and fully mimics the effects of natural and engineered mutations seen in full mouse models. Data deposited here comprises chromatin profiling of the EB derived CD71+ erythroid cells; chromatin accessibility (ATAC-Seq), characterisation of the open chromatin by associated histone modifications (ChIP-Seq), chromatin structure (CTCF ChIP-Seq and Capture C interaction data). WT CD71+ cells were compared to either mutant CD71+ cells or other erythroid tissues (mouse adult spleen, E13.5 fetal liver, early embryonic circulating blood) representing both definitve and primitive erythroid lineages.
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| Overall design |
NG Capture-C combines 3C library preparation with oligonucleotide capture for the desired viewpoint restriction fragments (DpnII fragment), in this case the Alpha Globin R1 enhancer and promoters of Rhbdf1 and Mpg genes. Purified CD71+ erythroid cells derived from differentiated Wildtype (WT) or mutant mouse Embryonic Stem (mES), D3839, deleted for the CTCF boudary elements at HS-38 and HS-39.5 positions at the Alpha Globin locus (Hanssen L et al, Nature Genetics, 2017) were used in this assay. The profiles produced from WT and D3839 CD71+ cells were then compared.
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| Contributor(s) |
Kassouf M, Gosden M |
| Citation(s) |
34995303, 39863595 |
| |
| Submission date |
Sep 20, 2021 |
| Last update date |
Feb 19, 2025 |
| Contact name |
Mira Tony Kassouf |
| E-mail(s) |
mira.kassouf@imm.ox.ac.uk
|
| Organization name |
University of Oxford
|
| Department |
Weatherall Institute of Molecular Medicine
|
| Lab |
Doug Higgs
|
| Street address |
John Radcliffe Hospital, Headly way
|
| City |
Oxford |
| State/province |
England |
| ZIP/Postal code |
OX3 9DS |
| Country |
United Kingdom |
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| Platforms (2) |
| GPL16417 |
Illumina MiSeq (Mus musculus) |
| GPL19057 |
Illumina NextSeq 500 (Mus musculus) |
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| Samples (12)
|
| GSM5588950 |
WT EBd7 CD71+ R1 Capture sample 1(WT_EB8_Enhancers) |
| GSM5588951 |
WT EBd7 CD71+ R1 Capture sample 2 (WT_EB9_Enhancers) |
| GSM5588952 |
WT EBd7 CD71+ R1 Capture sample 3 (WT_EB11_Enhancers) |
| GSM5588953 |
WT EBd7 CD71+ Rhbdf1 and Mpg Captures sample 1(WT_EB8_5Promoters) |
| GSM5588954 |
WT EBd7 CD71+ Rhbdf1 and Mpg Captures sample 2 (WT_EB9_5Promoters) |
| GSM5588955 |
WT EBd7 CD71+ Rhbdf1 and Mpg Captures sample 3 (WT_EB11_5Promoters) |
| GSM5588956 |
D3839 EBd7 CD71+ R1 Capture sample 1 (CTCFDKO_A3_Enhancers) |
| GSM5588957 |
D3839 EBd7 CD71+ R1 Capture sample 2 (CTCFDKO_A81_Enhancers) |
| GSM5588958 |
D3839 EBd7 CD71+ R1 Capture sample 3 (CTCFDKO_A82_Enhancers) |
| GSM5588959 |
D3839 EBd7 CD71+ Rhbdf1 and Mpg Captures sample 1 (CTCFDKO_A3_5Promoters) |
| GSM5588960 |
D3839 EBd7 CD71+ Rhbdf1 and Mpg Captures sample 2 (CTCFDKO_A81_5Promoters) |
| GSM5588961 |
D3839 EBd7 CD71+ Rhbdf1 and Mpg Captures sample 3 (CTCFDKO_A82_5Promoters) |
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| This SubSeries is part of SuperSeries: |
| GSE184435 |
Scalable Production of Mouse Erythroid Cells using an in vitro Embryoid Body differentiation System |
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| Relations |
| BioProject |
PRJNA764670 |
| SRA |
SRP337859 |
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