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| Status |
Public on Sep 11, 2020 |
| Title |
mESC2 |
| Sample type |
SRA |
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| Source name |
mESC_Untreated
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| Organism |
Mus musculus |
| Characteristics |
cell type: Embryonic Stem Cells
treatment: Untreated
ribo-depletion: Ribo-Zero
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| Growth protocol |
E14 mESCs were grown on 0.1% gelatin coated plates in 2i medium36 containing: DMEM/F12 (Gibco, 31331) and Neurobasal (Gibco, 12348) 1:1, N2 supplement (Gibco, 17502048), B27 supplement (Gibco, 17504044), 1X glutaMax (Gibco, 35050061), 1X penicilin/streptomycin (Gibco, 15140122), 1 mM sodium pyruvate (Gibco, 11360070), 50 nM 2-mercaptoethanol (Gibco, 31350010), nonessential amino acids (Gibco, 11140076), LIF, 3 μM GSK3 inhibitor (CHIR-99021) and 1 μM MEK inhibitor (PD0325901). They were differentiated into neurons as previously described.
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| Extracted molecule |
total RNA |
| Extraction protocol |
20 µg TriZOL purified RNA from each sample was depleted of rRNA using a Ribo-Zero rRNA Magnetic Kit (Epicentre) including the optional RiboGuard RNase inhibitor according to manufacturer’s protocol. The concentration was normalised so that each sample contained the same amount of RNA. To 1/3 of the sample 1/10 of the recommended amount of spike-in (ERCC RNA spike-in mix, Ambion) was added, ethanol precipitated, and resuspended in ‘Elute, fragment, finish mix’ (Illumina). The remaining 2/3 of the sample was ethanol precipitated and resuspended in 15 µl nuclease free water. The sample was heated to 70˚C for 1 min and incubated on ice for 2 min. 5 µl RNase R mixture (Epicentre) was added to the sample before incubation at 37˚C for 30 min. RNase R was removed by phenol/chloroform extraction. The RNA was resuspended in ‘Elute, fragment, finish mix’ (Illumina).
Sequencing libraries were prepared using Truseq stranded RNA LT kit (Illumina) from both Ribo-Zero and Ribo-Zero/RNase R samples, by fragmentation, 1st and 2nd strand cDNA synthesis, 3’-end adenylation, ligation of adaptors, and enrichment of DNA fragments using the manufacturer’s protocol
The samples were sequenced using the Illumina Illumina HiSeq 2500 platform with 100 bp paired-end reads (AROS Applied Biotechnology)
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| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina HiSeq 2500 |
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| Description |
mESC2_GTGAAA_L001
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| Data processing |
circRNA prediction was conducted using find_circ, ciri2 (v2.0.6) and circExplorer2 (v2.3) using default settings
Genome_build: mm10
Supplementary_files_format_and_content: Merged raw counts provided by ciri2, find_circ or circexplorer2
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| Submission date |
Sep 10, 2020 |
| Last update date |
Sep 11, 2020 |
| Contact name |
Thomas B Hansen |
| E-mail(s) |
tbh@mbg.au.dk
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| Phone |
20733619
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| Organization name |
MBG, Aarhus University
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| Street address |
C.F. Moellers Alle 3, build 1130
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| City |
Aarhus C |
| State/province |
State... |
| ZIP/Postal code |
8000 |
| Country |
Denmark |
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| Platform ID |
GPL17021 |
| Series (1) |
| GSE157788 |
circZNF827 nucleates a transcription inhibitory complex to balance neuronal differentiation |
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| Relations |
| BioSample |
SAMN16093819 |
| SRA |
SRX9104645 |
| 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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