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Sample GSM2509598 Query DataSets for GSM2509598
Status Public on Jun 13, 2017
Title Pcgf1KO+ Pcgf1_flag RNAseq
Sample type SRA
Source name Pcgf1KO+ Pcgf1_flag, mESCs
Organism Mus musculus
Characteristics strain: C57BL/6
cell type: mouse embryonic stem cells
genotype: Pcgf1KO+ Pcgf1_flag
Treatment protocol Pcgf1 KO ES cells were generated by Cas9 technology. Pcgf1KO+Pcgf1_flag ES cells were established by lentiviral infection technology.
Growth protocol ES cells were cultured with mouse embryonic fibroblasts (MEFs) inactivated with mitomycin-C in DMEM supplemented with15% fetal calf serum , leukemia inhibitory factor , L-glutamine , non-essential amino acids , 0.1 mM betamercaptoethanol , and penicillin/streptomycin. All ES cell lines were maintained at 37°C in 5% CO2.
Extracted molecule total RNA
Extraction protocol RNA was prepared using the TRIzol reagent following the manufacturer's recommendations. RNA was quantified using a NanoDrop-1000 spectrophotometer.
A total amount of 2 μg RNA per sample was used as input material for the RNA sample preparations. Sequencing libraries were generated using NEBNext® Ultra™ RNA Library Prep Kit for Illumina® (#E7530L, NEB, USA) following the manufacturer’s recommendations and index codes were added to attribute sequences to each sample. Briefly, mRNA was purified from total RNA using poly-T oligo-attached magnetic beads. Fragmentation was carried out using divalent cations under elevated temperature in NEBNext First Strand Synthesis Reaction Buffer (5X). First strand cDNA was synthesized using random hexamer primer and RNase H. Second strand cDNA synthesis was subsequently performed using buffer, dNTPs, DNA polymerase I and RNase H. The library fragments were purified with QiaQuick PCR kits and elution with EB buffer, then terminal repair、A-tailing and adapter added were implemented. The aimed products were retrieved by agarose gel electrophoresis and PCR was performed, then the library was completed.
Library strategy RNA-Seq
Library source transcriptomic
Library selection cDNA
Instrument model HiSeq X Ten
Data processing Mapping reads with the reference genome use TopHat 2.0.12
Assemble reads use Cufflinks 2.2.1
Gene expression level analysis HTSeq 0.6.0
Alternative splicing prediction use Cuffcompare 2.2.1
Differential expression gene analysis use DEGSeq 1.18.0
Genome_build: mm10
Supplementary_files_format_and_content: tab-delimited text files include FPKM values for each Sample …
Submission date Feb 26, 2017
Last update date May 15, 2019
Contact name Wukui Zhao
Phone 86-02558641524
Organization name Nanjing University
Department Model Animal Research Center
Lab MOE Key Laboratory of Model Animal for Disease Study
Street address 12 Xuefu Road
City Nanjing
State/province Jiangsu
ZIP/Postal code 210061
Country China
Platform ID GPL21273
Series (1)
GSE95383 Expression regulation by Pcgf1 in mouse embryonic stem cells
BioSample SAMN06335400
SRA SRX2577113

Supplementary file Size Download File type/resource
GSM2509598_Pcgf1KO+_Pcgf1_flag.txt.gz 2.6 Mb (ftp)(http) TXT
SRA Run SelectorHelp
Raw data are available in SRA
Processed data provided as supplementary file

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