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Sample GSM1100743 Query DataSets for GSM1100743
Status Public on Sep 13, 2013
Title NPC_ko_rep1
Sample type SRA
Source name Neural progenitor cells
Organism Mus musculus
Characteristics cell type: NPC
treatment: Prmt5 depletion
developmental stage: E14.5
genotype: Prmt5 F/FER
treatment: 4-OHT
strain: mixed C57BL/6 X 129S1/SvlmJ
Treatment protocol PRMT5F/F ER day 4 NPCs and MEFs were treated with either 50nM 4-OHT or the equivalent volume of ethanol for 24 hours before splitting to induce PRMT5 knockout.
Growth protocol Neurosphere cultures were established as previously described. Briefly, E14.5 embryos were harvested and cortices carefully dissected in ice-cold PBS and incubated in trypsinfor 10 min at 37°C. The tissue was then mechanically dissociated into single cell suspension and passed through a 40 μm cell strainer into complete NSC medium, 1% penicillin-streptomycin, 20 ng/ml recombinant human epidermal growth factor and 20 ng/ml recombinant human fibroblast growth factor-basic. Mouse Embryonic Fibroblasts (MEFs). Primary MEFs were prepared from E14.5 embryos as previously described 48 and maintained in a humidified 5% CO2 atmosphere at 37°C in DMEM supplemented with 10% fetal bovine serum and 1% penicillin-streptomycin. To induce PRMT5 knockout, MEFs (passage 1) were grown to confluence in 15 cm-dishes.
Extracted molecule total RNA
Extraction protocol RNA was harvested using Trizol reagent.
Illumina TruSeq RNA Sample Prep Kit v2(Cat#FC-122-1001) was used with 1 ug of total RNA for the construction of sequencing libraries.
Library strategy RNA-Seq
Library source transcriptomic
Library selection cDNA
Instrument model Illumina HiSeq 2000
Data processing Illumina Casava v1.8 used for basecalling.
Sequences were mapped to mm9 using TopHat v2.03 and Bowtie with parameters -m 1 -n 2 --no-coverage-search --library-type fr-unstranded --solexa-quals
FPKM values were computed using cuffdiff 2.02 with parameters -u --total-hits-norm -N
To determine differential splicing events, MATS 3.0.6 beta was used counting junction reads and reads falling into the tested region within ENSEMBL v65 gene definitions. Matching embryos were analysed individually and only significant events occurring in at least two replicates were considered. Splicing events were labelled significant if the sum of the reads supporting a specific event exceeded 10 reads, the p-value was lower than 0.05 and the minimum inclusion level difference as determined by MATS was higher than 0.2. All other parameters were left at the default value.
Genome_build: mm9
Supplementary_files_format_and_content: tab-delimited text files include FPKM values and differential splicing results for each sample
Submission date Mar 19, 2013
Last update date May 15, 2019
Contact name Julius Müller
Organization name A*Star Singapore
Department IMCB
Lab Ernesto Guccione
Street address 61 Biopolis Drive
City Singapore
ZIP/Postal code 138673
Country Singapore
Platform ID GPL13112
Series (2)
GSE45284 Regulation of constitutive and alternative splicing by PRMT5 reveals a role for Mdm4 pre-mRNA in sensing defects in the spliceosomal machinery (RNA-Seq)
GSE45285 Regulation of constitutive and alternative splicing by PRMT5 reveals a role for Mdm4 pre-mRNA in sensing defects in the spliceosomal machinery
SRA SRX252191
BioSample SAMN01984502

Supplementary data files not provided
SRA Run SelectorHelp
Raw data are available in SRA
Processed data are available on Series record

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