 |
 |
GEO help: Mouse over screen elements for information. |
|
| Status |
Public on Dec 31, 2017 |
| Title |
MeDIP_Hippocampal_neurons_4h_KCL_rep2 |
| Sample type |
SRA |
| |
|
| Source name |
hippocampal neurons
|
| Organism |
Mus musculus |
| Characteristics |
tissue: cultured hippocampal neurons
differentiation state in vitro: 11 days
developmental stage: E18.5
strain: C57BL/6J
|
| Treatment protocol |
samples treated or untreated with 55mM KCl
|
| Growth protocol |
1000Xg, cells were resuspended in complete NB medium containing 22µM L-glutamic acid (Sigma, Germany), counted, and seeded on poly-ornithine (0.1 mg/ml, Sigma) and laminin (1μg/ml, Sigma) coated plates. Medium change was performed every 96h by exchanging half of the volume of the medium with fresh NB medium with complements and AraC (1µg/ml for the first, and 0.5µg/ml for the second medium change, Sigma).
|
| Extracted molecule |
genomic DNA |
| Extraction protocol |
C57BL/6J (Janvier Labs, France) hippocampi of E18.5 embryos were dissected and collected in 15 ml Hanks’ Balanced Salt Solution (HBSS, PAA, Germany). After centrifugation (5min; 800xg) hippocampi were dissociated by adding 1ml 0.05% trypsin-ethylenediaminetetracetic acid (PAA) at 37°C for 15min. After adding 1ml of fetal bovine serum (FBS, Life Technologies) and 1ml of neurobasal (NB) medium supplemented as previously described (Vezzali R et al. 2016) samples were homogenized by repeated passages through a pipette tip. After 10min centrifugation at 1000Xg, cells were resuspended in complete NB medium containing 22µM L-glutamic acid (Sigma, Germany), counted, and seeded on poly-ornithine (0.1 mg/ml, Sigma) and laminin (1μg/ml, Sigma) coated plates. Medium change was performed every 96h by exchanging half of the volume of the medium with fresh NB medium with complements and AraC (1µg/ml for the first, and 0.5µg/ml for the second medium change, Sigma). At day in vitro (DIV) 11 cells were treated with 55mM KCl at 37ºC, 5%CO2 at different time points before harvesting.
The company Active motif processed the samples, constructed the library and analyzed the data.
|
| |
|
| Library strategy |
MeDIP-Seq |
| Library source |
genomic |
| Library selection |
5-methylcytidine antibody |
| Instrument model |
Illumina NextSeq 500 |
| |
|
| Description |
Name of company: 4_2577Albert_Hippo_KCl_5mC_i89
Hippocampal neurons were isolated at day E18.5, cultured for 11 days in vitro and then treated with or without 55mM KCL, then MeDIP-seq was performed
|
| Data processing |
The sequence reads identified by the Active Motif Sequencing Service (NextSeq 500, 75-nt single-end, flowcell AH1350BGXX) are mapped to the genome using the BWA algorithm with default settings.Alignment information for each read is stored in the BAM format. Only reads that pass Illumina’s purity filter, align with no more than 2 mismatches, and map uniquely to the genome are used in the subsequent analysis. In addition, unless stated otherwise, duplicate reads (“PCR duplicates”) are removed.
Since the 5´-ends of the aligned reads (= “tags”) represent the end of ChIP/IP-fragments, the tags are extended in silico (using Active Motif software) at their 3´- ends to a length of 150-250 bp, depending on the average fragment length in the size selected library (normally 150 bp). To identify the density of fragments (extended tags) along the genome, the genome is divided into 32-nt bins and the number of fragments in each bin is determined.
The two main peak callers used at Active Motif are MACS (Zhang et al., Genome Biology 2008, 9:R137) and SICER (Zang et al., Bioinformatics 25, 1952-1958, 2009).
To compare peak metrics between 2 or more samples, overlapping Intervals are grouped into “Active Regions”, which are defined by the start coordinate of the most upstream Interval and the end coordinate of the most downstream Interval (= union of overlapping Intervals). In locations where only one sample has an Interval, this Interval defines the Active Region.
Genome_build: mm10
|
| |
|
| Submission date |
Nov 25, 2016 |
| Last update date |
May 15, 2019 |
| Contact name |
Henriette Franz |
| Organization name |
University Freiburg
|
| Department |
Anatomy and Cellbiology
|
| Lab |
Vogel
|
| Street address |
Alberstr: 17
|
| City |
Freiburg |
| ZIP/Postal code |
79104 |
| Country |
Germany |
| |
|
| Platform ID |
GPL19057 |
| Series (2) |
| GSE90509 |
Neuronal activity, TGFβ signaling and unpredictable chronic stress affect transcription of Gadd45 family members in vitro and in vivo [MeDIP-Seq] |
| GSE90513 |
Neuronal activity, TGFβ signaling and unpredictable chronic stress affect transcription of Gadd45 family members in vitro and in vivo |
|
| Relations |
| BioSample |
SAMN06054315 |
| SRA |
SRX2375297 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSM2405860_MeDIP_HN_4h_KCL_rep2.bw |
65.9 Mb |
(ftp)(http) |
BW |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data provided as supplementary file |
|
|
|
|
 |
