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| Status |
Public on Jan 22, 2025 |
| Title |
p300_iHAT_NT_rep3_Donor_A486_2 |
| Sample type |
SRA |
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| Source name |
endoderm_stage18
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| Organism |
Xenopus laevis |
| Characteristics |
tissue: endoderm_stage18
cell type: endoderm
genotype: WT
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| Treatment protocol |
IVF-derived gastrulae (NF stage 12) were treated with 40 µM SGC-CBP30 (Sigma, SML1133), 30 µM A-485 (Tcris, #6387) or DMSO in 0.1x MMR at 23°C until they reached neurula stage (NF stage 18)59. The embryos were collected at synchronized stages to eliminate influences from different cell numbers or developmental stages. The embryos were either used as donors for nuclear transfer as described above, or frozen for RNA-extraction.Endoderm was dissected from neurula-stage embryos (NF stage 18) and allowed to dissociate into single cells in calcium- and magnesium-free 1x modified Barth saline (MBS, 88 mM NaCl, 1mM KCl, 10mM HEPES, 2.5 mM NaHCO3, pH 7.4) with 1 mM EDTA and 0.1% BSA in a Petri dish coated with 0.1% agarose in H2O. Donor cells were immediately used for NT.
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| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA was isolated using the RNeasy Mini Kit (QIAGEN, 74104) according to the manufacturer’s instructions. To lyse the embryonic tissue, samples were vortexed on high speed for 10 min at 4°C. DNase digestion was performed according to the manufacturer’s instructions, using RNase-free DNase (QIAGEN, 79254).
Per sample, 400 ng total RNA of animal cap (ectoderm) tissue and 300 ng of endoderm donor tissue were used to isolate mRNA using the NEBNext Poly(A) mRNA Magnetic Isolation Module (NEB, E7490). Sequencing libraries were generated using the NEBNext Ultra II Directional RNA Library Prep Kit for Illumina (NEB, E7760), following the manufacturer’s instructions, using 12-13 PCR amplification cycles. Libraries were multiplexed and sequenced on a NextSeq 6000 platform.
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| Library strategy |
RNA-Seq |
| Library source |
transcriptomic |
| Library selection |
cDNA |
| Instrument model |
Illumina NovaSeq 6000 |
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| Description |
Library name: p300_iHAT_NT_rep3_Donor_A486_2
2x50 bp reads
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| Data processing |
Paired sequencing reads were processed using Kallisto (v0.48) for pseudoalignment and quantification of transcript abundance.
Transcript and annotation files were downloaded from Xenbase (v10.1) for Xenopus laevis (transcripts and annotation).
After quantification, transcript-level abundances were imported using 'tximport' and converted to a 'SummarizedExperiment' object in R (v4.3.1).
Datasets of two independent batches were merged and subjected to differential expression analysis performed with DESeq2 (v1.40.2).
Assembly: Xenopus laevis 10.1
Supplementary files format and content: Kallisto output containing transcript abundance as transcript per million.
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| Submission date |
Dec 22, 2024 |
| Last update date |
Jan 22, 2025 |
| Contact name |
Eva Hoermanseder |
| E-mail(s) |
eva.hoermanseder@helmholtz-munich.de, antonio.scialdone@helmholtz-munich.de
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| Organization name |
Helmholtz Center Munich
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| Department |
Institute of Epigenetics and Stem Cells
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| Lab |
Hoermanseder Lab
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| Street address |
Feodor-Lynen-Str. 21
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| City |
Munich |
| State/province |
Bavaria |
| ZIP/Postal code |
81377 |
| Country |
Germany |
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| Platform ID |
GPL28901 |
| Series (1) |
| GSE285204 |
‘Digital Reprogramming’ Decodes Epigenetic Barriers of Cell Fate Changes and Identifies p300 Inhibitors as Facilitators [RNA-Seq] |
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| Relations |
| BioSample |
SAMN45942791 |
| SRA |
SRX27164656 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSM8698192_iH_hash4_A486_donor_hash2.abundance.tsv.gz |
785.3 Kb |
(ftp)(http) |
TSV |
SRA Run Selector |
| Raw data are available in SRA |
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