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GEO help: Mouse over screen elements for information. |
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| Status |
Public on Nov 19, 2018 |
| Title |
KO_8D_704507 |
| Sample type |
SRA |
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| Source name |
B cells
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| Organism |
Mus musculus |
| Characteristics |
tissue: bone marrow cell
cell type: B cells
tag: P198A/P198A mutant
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| Treatment protocol |
BM cells from straight chimeras were stained with B220,CD43,CD45.1,CD45.2,CD24,BP-1,IgM and IgD Abs.Cells were sorted using a FacsARIA III (BD), using the following gating strategy: viable, B220+CD43+CD45.1-CD45.2+CD24+BP1- (FrB/pro-B); B220+CD43-CD45.1-CD45.2+IgM-IgD- (FrD/late preB) and viable, B220+CD43-CD45.1-CD45.2+IgM+IgD- (FrE/Immature)
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| Growth protocol |
BM cells of WT and P198A/P198A mutant mice (generated by CRISPR/Cas9) were injected into lethally irradiated CD45.1 mice to generate straight chimeras. After 8 weeks reconstitution, 5 WT mice and 5 P198A/P198A mutant straight chimeras mice BM were freshly isolated for the experiment.
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| Extracted molecule |
polyA RNA |
| Extraction protocol |
100 cells per sample were directly sorted into ice cold cell lysis buffer (0.4% (vol/vol) Triton X-100 and 2 U/μl RNase inhibitor, 4 x107 dilution of ERCC spike in control, comprising a pre-formulated blends of 92 transcripts, derived and traceable from NIST-certified DNA plasmids), 2.5mM dNTPs (Thermo-Fisher), 2.5µM Oligo (Oligo-dT30VN.) and immediately frozen in dry ice.Sorted cells were processed using the Smart-seq2 protocol with minor changes
Indexing of the samples was performed using the Nextera XT DNA Sample Preparation Index kit (24 index primers, manufacturer (Illumina)), according to manufacturer’s instructions.
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| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina HiSeq 4000 |
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| Data processing |
Bioinformatic analysis was performed using CGAT Pipelines and tools (https://github.com/CGATOxford/CGATPipelines). Reads were aligned to the mm10 mouse genome using hisat2 version 2.1.0, and reads were quantified over feature annotations (ensemble81) using featureCounts program version 1.4.6 within the Subread software package. DESeq2 was used for statistical analysis of the differential expression for each gene between wild-type and ZIP7P198A/P198A mutants.
Genome_build: mm10
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| Submission date |
Dec 18, 2017 |
| Last update date |
Nov 19, 2018 |
| Contact name |
Adam Cribbs |
| E-mail(s) |
[email protected]
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| Organization name |
University of Oxford
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| Department |
NDORMS
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| Street address |
Windmill Road
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| City |
Oxford |
| ZIP/Postal code |
OX37LD |
| Country |
United Kingdom |
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| Platform ID |
GPL21103 |
| Series (1) |
| GSE108178 |
Human immunodeficiency reveals the Zn2+-dependence of B cell development |
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| Relations |
| BioSample |
SAMN08196410 |
| SRA |
SRX3478227 |
| Supplementary data files not provided |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data are available on Series record |
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