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Sample GSM2692017 Query DataSets for GSM2692017
Status Public on Jan 25, 2018
Title KO-biological replicate 1-technical replicate 2
Sample type SRA
Source name bone marrow derived mast cell
Organism Mus musculus
Characteristics cell type: bone marrow derived mast cell
strain: C57BL/6
genotype: Tet2-/-
Growth protocol Mouse bone marrow cells were isolated from femurs and cultured in RPMI1640 medium plus 10% (vol/vol) FBS (Gibco) with 10 ng/ml IL-3 and 5 ng/ml SCF. Half of the medium was replaced by fresh medium with cytokines every 2–3 d during culture. After 4–6 wk in culture, BMMCs were stained to confirm the surface expression of FcɛRI and c-Kit. Cells with purity >97.5% were used for subsequent experiments.
Extracted molecule polyA RNA
Extraction protocol Total RNAs were isolated by Trizol reagent. Poly-A tailed RNAs were enriched by GenElute™ mRNA Miniprep Kit (MRN70-1KT, Sigma). The mRNAs were treated with TURBO DNase, purified by a RNeasy MinElute Cleanup Kit (QIAGEN). Treated mRNAs with 0.5% unmethylated RNA controls from lambda genome were bisulfite converted using the EZ RNA Methylation kit (R5001, Zymo Research). Briefly, 500 ng mRNA was converted using three cycles of 10-min denaturation at 70℃ followed by 45min at 54℃. RNA separation from bisulfite solution, desulfonation, and purification were performed following the standard protocol of the kit.
Strand-specific libraries were constructed following the “TruSeq stranded mRNA sample preparation guide” from illumina for 150PE sequencing using Hiseq3000.
Library strategy Bisulfite-Seq
Library source transcriptomic
Library selection RANDOM
Instrument model HiSeq X Ten
Description FcɛRI+ c-Kit+, purity >97.5%
Data processing Illumina Casava1.6 software used for basecalling.
Reads mapping and methylcytosine calling were analyzed by BS-RNA using converted mm10 and lambda genome as references.
BS-RNA: An efficient mapping and annotation tool for RNA bisulfite sequencing data. Computational biology and chemistry, doi:10.1016/j.compbiolchem.2016.09.003 (2016).
Splice sites information was added using refGene.gtf file during read mapping.
Cs with methylation rate above 0.1 and at least two reads supporting methylation in both technical and biological replicates of one group and below the non-conversion rate of each of the replicates of the other group (WT1-rep1:0.0015, WT1-rep2:0.0015, KO1-rep1:0.0017, KO1-rep2:0.0016, WT2-rep1:0.0012, WT2-rep2:0.0013, KO2-rep1:0.0012, KO2-rep2:0.0012,) were chosen as group-specific Cs.
Genome_build: GRCm38/mm10
Submission date Jul 03, 2017
Last update date May 15, 2019
Contact name Qian Zhang
Phone +8615921196443
Organization name Second military medical university
Street address xiangyin road 800
City shang hai
ZIP/Postal code 200433
Country China
Platform ID GPL21273
Series (1)
GSE100719 Distinct 5-methylcytosine profiles in poly(A)RNA from mouse bone marrow derived mast cells
BioSample SAMN07311769
SRA SRX2978628

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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