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Sample GSM4431941 Query DataSets for GSM4431941
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 SelectorHelp
Raw data are available in SRA
Processed data provided as supplementary file

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