 |
 |
GEO help: Mouse over screen elements for information. |
|
| Status |
Public on Jul 27, 2017 |
| Title |
1835_cortical_d27 |
| Sample type |
SRA |
| |
|
| Source name |
Cortical neuron stem cells differentiated from iPSC
|
| Organism |
Homo sapiens |
| Characteristics |
developmental stage: Cortical NSC/neurons at day 27 of neuronal differentiation
Sex: female
passage number: 22
|
| Treatment protocol |
NA
|
| Growth protocol |
For iPSC culture and cortical neuron differentiation, we have adopted the protocol (Shi et al., Nature Protocols, 2012) to our iPSCs growing In feeder-free medium (mTeSR). In brief, we plate 5 wells of iPSCs (on a 6-well plate) onto one well of Geltrax-coated 12-well plate in 800ul of mTeSR media, allowing cells to attach overnight and reaching 100% confluence 1 day after plating. For neural induction, we switched the cell culture media to neural induction media (NIM) containing 1uM of Dorsomorphin. After 10 days of neural induction when a homogeneous neuroepithelial layer is formed, we passaged the cells onto laminin-coated 35-mm dish and switched the cell culture media to Neural Maintenance Medium (NMM). Upon appearance of neural rosettes, we added 20ng/ml of FGF2 for 2-4 days to promote the expansion of neural stem cells (NSCs). At about day 20 after neural induction, we passaged the cells for the first NSC expansion (1: 3). After another round of NSC expansion at day 25, we passaged the NSC culture at ~day 30 using Accutase at a ratio of 1:4 onto laminin-coated 35-mm dish (for collecting cells later) and laminin-coated coverslips (for immunofluorescence staining). We continued the neural differentiation for the plated cells in NMM until the specified days to collect the cells.
|
| Extracted molecule |
genomic DNA |
| Extraction protocol |
For ATAC-seq, we harvested 50,000 cells (iPSCs, induced NSCs and neurons at days 27, 33 and 41 of neural induction) by centrifugation at 500g x 5 min at 4°C. We washed cells once with PBS buffer and resuspended the cell pellet in 50 μL of cold lysis buffer (10 mM Tris-HCl, pH7.4, 10 mM NaCl, 3 mM MgCl2, 0.1% IGEPAL CA-630). We spinned down cell pellet, discarded the supernatant, and immediately continued to transposition reaction (keep the nuclei pellet on ice) in 50 μl transposition reaction mix. We incubated the transposition reaction at 37°C for 30 min, immediately followed by purification through a Qiagen MinElute Kit. We stored purified DNA at -20°C until library creating sequencing library.
The library was generated and sequenced at University of Minnesota Genomic Center. We have followed the protocol as previously described (Buenrostro et al., Nature Methods. doi:10.1038/nmeth.2688, 2013) except for using primers from the Illumina Nextera kit for PCR amplification of the transposed DNA fragments.
|
| |
|
| Library strategy |
OTHER |
| Library source |
genomic |
| Library selection |
other |
| Instrument model |
Illumina HiSeq 2500 |
| |
|
| Description |
day 27 culture; but was considered as duplicate of day 30 because of high correlation with day 33 sample
|
| Data processing |
Library strategy: ATAC-Seq
Library strategy: ATAC-Seq
Illumina CASAVA v1.8 standard
Reads were trimmed for adaptor sequence, then mapped to UCSC hg19 using bowtie, duplicate fragments were then removed using Picard.
Genome-wide read density was calculated as the number of insertions found within 150bp sliding window with 20 bp step size.
Peaks were called using the HOTSPOT v4 with FDR set to 0.05 and the default settings. The 50bp mappability files were downloaded from http://www.uwencode.org/proj/hotspot/.
Genome_build: hg19
Supplementary_files_format_and_content: The hotspot peaks are stored in a tab delimited bed file with column 1 indicating the chromosome column 2 and 3 indicate the start and end of the peak. The scores stored in the 5th columnare z-scores that are from the binomial model for scoring hotspots
|
| |
|
| Submission date |
Jul 13, 2015 |
| Last update date |
May 15, 2019 |
| Contact name |
Winton Moy |
| E-mail(s) |
winton.moy81@gmail.com
|
| Phone |
224-364-7567
|
| Organization name |
Northshroe Univeristy Health Systems
|
| Department |
Center Psychiatric Genetics
|
| Street address |
1001 University Place
|
| City |
Evanston |
| State/province |
Illinois |
| ZIP/Postal code |
60201 |
| Country |
USA |
| |
|
| Platform ID |
GPL16791 |
| Series (1) |
| GSE70823 |
Mapping open chromatin dynamics of cortical neuron differentiation from human induced pluripotent stem cells (iPSCs) by ATAC-Seq |
|
| Relations |
| BioSample |
SAMN03856503 |
| SRA |
SRX1092537 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSM1820082_d27_fdr0.05.hot.bed.gz |
1.2 Mb |
(ftp)(http) |
BED |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data provided as supplementary file |
|
|
|
|
 |
