 |
 |
GEO help: Mouse over screen elements for information. |
|
Status |
Public on Sep 04, 2020 |
Title |
Input replicate 3 |
Sample type |
SRA |
|
|
Source name |
Skin epithelial cells
|
Organism |
Mus musculus |
Characteristics |
strain: C57BL/6 tissue: Skin age: Postnatal day 0 genotype: wild-type
|
Extracted molecule |
polyA RNA |
Extraction protocol |
Back skin dissected from P0 neonates was treated first with 1x dispase to isolate epidermis, which was then digested with 1:1 0.25% Trypsin-EDTA:versene solution to break down tissue. Single cell suspensions were obtained by filtering through 45 µm cell strainers. Epithelial progenitor cells were isolated by FACS. Total RNA was extracted by TRIzol-LS. Poly(A)+ RNA was extracted with the Dynabeads™ mRNA Purification Kit (Thermo-Fisher, 61006) and treated with the RQ1 RNase-free DNase (Promega, M6101) to remove DNA contamination. 7-8 µg Poly(A)+ RNA was used for each biological replicate (pool of 3 litters of pups) to perform miCLIP. After fragmentation, 1/10 of the sample was saved as input to perform parallel library construction without CLIP. Library construction protocol is as described in Geula et al., 2015, Mapping m6A at Individual-Nucleotide Resolution Using Crosslinking and Immunoprecipitation (miCLIP)
|
|
|
Library strategy |
RIP-Seq |
Library source |
transcriptomic |
Library selection |
other |
Instrument model |
Illumina NextSeq 500 |
|
|
Description |
Poly(A)+ RNA
|
Data processing |
Fastq data was adaptor and quality trimmed using Flexbar with options (—pre-trim-phred 30, -n 20, -s) Trimmed Fastq was demultiplexed using pyCRAC’s pyBarcodeFilter.py script and deduplicated using the pyFastqDuplicateRemover.py script Fastq was aligned to the mm10 genome using Novoalign with options (-t 85, -l 16, -s 1, -o Native, -r None, -c 20, -a). Aligned reads were converted to novoalign2bed.pl script in the CIMS CTK toolkit. m6A sites were called using the CIMS pipeline as part of the CTK toolkit with default parameters Genome_build: Mm10 (GRCm38 - Dec. 2011) genome build Supplementary_files_format_and_content: Bigwigs were generated from aligned reads as BED format using the GenomicRanges R package and raw coverage generated by rtracklayer R package.
|
|
|
Submission date |
Mar 24, 2020 |
Last update date |
Sep 04, 2020 |
Contact name |
Tom Samuel Carroll |
E-mail(s) |
thomas.carroll@rockefeller.edu
|
Organization name |
The Rockefeller University
|
Department |
Bioinformatics
|
Lab |
Bioinformatics
|
Street address |
1230 York Avenue
|
City |
New York |
State/province |
New York |
ZIP/Postal code |
10065 |
Country |
USA |
|
|
Platform ID |
GPL19057 |
Series (2) |
GSE147489 |
miCLIP-seq of postnatal day 0 (P0) normal mouse skin epithelial cells |
GSE147490 |
mouse skin epithelial cells |
|
Relations |
BioSample |
SAMN14442116 |
SRA |
SRX7988129 |
Supplementary file |
Size |
Download |
File type/resource |
GSM4431941_Rep3In_tag_neg.bigwig |
50.8 Mb |
(ftp)(http) |
BIGWIG |
GSM4431941_Rep3In_tag_pos.bigwig |
51.0 Mb |
(ftp)(http) |
BIGWIG |
SRA Run Selector |
Raw data are available in SRA |
Processed data provided as supplementary file |
|
|
|
|
 |