|
| Status |
Public on Jun 23, 2015 |
| Title |
Adelman_Bl6_MEF_startRNA-seq_rep1 |
| Sample type |
SRA |
| |
|
| Source name |
Adelman_Bl6_MEF_startRNA-seq
|
| Organism |
Mus musculus |
| Characteristics |
strain background: C57BL/6
gender: female
age: 8-12 weeks
cell type: mouse embryonic fibroblasts
treated with: none (untreated)
molecule subtype: short capped RNA isolated from MEF nuclei
|
| Treatment protocol |
BMDMs were incubated with 0 or 100ng/ml LPS (Sigma L6529) for the time indicated.
|
| Growth protocol |
For each biological replicate, macrophages were derived from female C57BL/6 mice (age 8-12 weeks). Day 7 bone marrow-derived macrophages were incubated at 37°C with 5% CO2. MEFs were derived from e13.5 female C57BL/6 embryos as follows: body tissue from e13.5 embryos was minced and incubated at 37°C with 0.5% Trypsin, followed by quenching with DMEM (10% FBS, 1X Penn/Strep, 1X Glutamine), and passage through a 100 micron mesh filter. Cells in suspension were plated, incubated at 37°C with 5% CO2, and passaged as necessary.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA was extracted from nuclei using Trizol reagent (Invitrogen)
Start-RNA libraries were prepared as described in (PMID:20007866) except reads were size selected in the range 20-80 nt to exclude full length snRNA species.
|
| |
|
| Library strategy |
ncRNA-Seq |
| Library source |
transcriptomic |
| Library selection |
size fractionation |
| Instrument model |
Illumina MiSeq |
| |
|
| Description |
processed data file:
Adelman_Bl6_MEF_startRNA-seq_5pr_rep1and2_forward.bedgraph.gz
Adelman_Bl6_MEF_startRNA-seq_5pr_rep1and2_reverse.bedgraph.gz
|
| Data processing |
Base calling and generation of FASTQ files performed by MiSeq Control Software 2.2.0 (MEF samples), standard Illumina GAIIx pipeline (macrophage samples)
Reads trimmed for adapter sequence using cutadapt 1.2.1, pairs containing reads shorter than 15 nt excluded (-q 10 -m 15, otherwise default), reads further trimmed to maximum length of 36 nt, MEF paired-end startRNA-seq only
Reads 3' trimmed to 36 nt (all macrophage data)
Reads aligned using bowtie 0.12.8 (-m1 -v2, -X1000 for paired-end alignments)
Technical and biological replicates merged
Counts normalized such that totals across all time points are equivalent (macrophage startRNA-seq, RNA PolII ChIP-seq)
Genome_build: mm9
Supplementary_files_format_and_content: bedgraph: 5' ends of uniquely aligned pairs (MEF samples); wig: normalized 5' ends of uniquely aligned single-end reads (macrophage samples), normalized center-shifted (65 nt), strand merged, and binned (25) RNA PolII ChIP-seq read counts; bigWig: 25 nt binned MNase-seq fragment centers
|
| |
|
| Submission date |
Oct 07, 2014 |
| Last update date |
May 15, 2019 |
| Contact name |
Karen Adelman |
| E-mail(s) |
karen_adelman@hms.harvard.edu
|
| Organization name |
Harvard Medical School
|
| Department |
Biological Chemistry and Molecular Pharmacology
|
| Street address |
45 Shattuck St. LHRRB-201a
|
| City |
Boston |
| State/province |
MA |
| ZIP/Postal code |
02115 |
| Country |
USA |
| |
|
| Platform ID |
GPL16417 |
| Series (1) |
| GSE62151 |
Upstream Anti-sense Promoters Act as Local Enhancers of Mammalian Protein-coding Genes |
|
| Relations |
| BioSample |
SAMN03098202 |
| SRA |
SRX727215 |
