|
| Status |
Public on Aug 31, 2018 |
| Title |
ESC_Total_nodox_3 (5) |
| Sample type |
SRA |
| |
|
| Source name |
embryonic stem cells
|
| Organism |
Mus musculus |
| Characteristics |
cells: ESC doxycycline-inducible expressing HA-Flag-AGO2
passages: 8-15
fraction: Total
rip antibody: Monoclonal ANTI-FLAG® M2 antibody produced in mouse (Sigma-Aldrich, catalog# M8823)
|
| Treatment protocol |
15 h before cell harvesting, transgene expression was induced with 2.5 ug/ml doxycycline in ESC.
|
| Growth protocol |
ESC were grown on 0.1% gelatin in DMEM supplemented with 15% heat-inactivated FBS, 0.1 mM 2-mercaptoethanol, 2 mM L-glutamine, 0.1 mM non-essential amino acids, 1mM Sodium Pyruvate and 1000 U/ml recombinant leukemia inhibitory factor.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Cells were fractionated and AGO2 was immunoprecipitated from cytoplasmic and nuclear fraction. The RNA was isolated using Trizol.
Preadenylated, barcoded 3´adapter oligonucleotide was ligated followed by ligation of a 5´adapter. Ligation products were reverse transcribed and amplified by PCR.
|
| |
|
| Library strategy |
miRNA-Seq |
| Library source |
transcriptomic |
| Library selection |
size fractionation |
| Instrument model |
Illumina HiSeq 3000 |
| |
|
| Description |
miRNA_ESC_MT.xlsx
|
| Data processing |
Illumina HiSeq 3000 platform was used for sequencing
Basecalling was done using bcl2fastq V2.17.1
The reads were aligned to the mm10 genome using TopHat
The miRNA normalized counts was generated as in Farazi et al., Cancer research 2011 (GEO accession number GSE28884)
Genome_build: mm10
Supplementary_files_format_and_content: xlsx reports Normalized counts
|
| |
|
| Submission date |
Jan 05, 2018 |
| Last update date |
Aug 31, 2018 |
| Contact name |
Markus Hafner |
| E-mail(s) |
Markus.hafner@nih.gov
|
| Phone |
3014026956
|
| Organization name |
NIH
|
| Department |
NIAMS
|
| Lab |
Markus Hafner
|
| Street address |
50 South Drive
|
| City |
Bethesda |
| State/province |
MD |
| ZIP/Postal code |
20892 |
| Country |
USA |
| |
|
| Platform ID |
GPL21493 |
| Series (2) |
| GSE108799 |
Nuclear miRNA-mediated gene silencing dependent on Argonaute proteins is required for embryonic stem cell differentiation [5_miRNA-seq] |
| GSE108801 |
Nuclear miRNA-mediated gene silencing dependent on Argonaute proteins is required for embryonic stem cell differentiation |
|
| Relations |
| BioSample |
SAMN08296067 |
| SRA |
SRX3534036 |
