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| Status |
Public on Dec 21, 2023 |
| Title |
PM_137_NPC_HiC |
| Sample type |
SRA |
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| Source name |
iPSC derived NPC
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| Organism |
Homo sapiens |
| Characteristics |
cell type: iPSC derived NPC
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| Growth protocol |
Human iPSC were differentiated into NPCs using a previously established protocol (Xie et al 2013). Briefly, undifferentiated cells were maintained in mTESR Plus (STEMCELL Technology, 05825) on Matrigel coated plates. They were seeded onto fresh Matrigel plates in NPC media at a density of 16,000 cells/cm2. NPC media was changed every day and cells were harvested at the end of day 8. The NPC differentiation medium consists of DMEM/F12 (Thermo Fisher, 11320033) with 5 μg/ml insulin, 64 μg/ml L-ascorbic acid, 14 ng/ml sodium selenite, 10.7 ug/ml Holo-transferrin, 543 μg/ml sodium bicarbonate, 10 μM SB431542 and 100 ng/ml Noggin.
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| Extracted molecule |
genomic DNA |
| Extraction protocol |
We prepared Hi-C libraries using the Arima Genomics Hi-C kit (Arima Genomics, A510008) according to the manufacturer’s protocol. Briefly, we enzymatically digested genomic DNA within nuclei of crosslinked cell pellets, and created biotinylated ligation junctions between the digested ends at proximity. Then we extracted DNAs and sheared to an average size of ~400 bp using a Covaris S220 sonicator at 140 W peak incident power, 10 % duty factor, and 200 cycles per burst for 55 seconds. We further size selected the sheared DNA to 200-600 bp using AgenCourt Ampure XP beads (Beckman Coulter, A63881) according to manufacturer’s protocols. Biotin-tagged ligation junctions via pulldown using streptavidin breads from the Arima Hi-C kit (Arima Genomics, A510008) according to manufacturer’s protocol. Streptavidin beads containing Hi-C libraries were stored at -20 °C for no more than 3 days before Illumina sequencing library preparation was performed.
Hi-C libraries were prepped for sequencing by first washing adaptor-ligated Hi-C libraries on streptavidin beads twice in 150 uL of wash buffer at 55 °C and once in 100 ml of elution buffer at room temperature using Hi-C kit (Arima Genomics, A510008). DNA was eluted from streptavidin beads by boiling at 98 °C for 10 min in 15 uL elution buffer. Subsequently the libraries were amplified using NEBNext Ultra II DNA Library Prep Kit for Illumina (NEB, E7645S) with 8 PCR cycles according to the manufacturer’s protocol.
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| Library strategy |
Hi-C |
| Library source |
genomic |
| Library selection |
other |
| Instrument model |
Illumina NextSeq 500 |
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| Data processing |
Paired-end reads were aligned independently to the hg38 human genome using bowtie2 (global parameters: --verysensitive –L 30 –score-min L,-0.6,-0.2 –end-to-end --reorder; local parameters: --very-sensitive –L 20 –score-min L,-0.6,-0.2 –end-to-end --reorder) through the HiC-Pro software8. Unmapped reads, non-uniquely mapped reads, and PCR duplicates were filtered and uniquely aligned reads were paired. Raw contact matrices for all samples were assembled into 20kb, 40kb, and 100kb non-overlapping bins and balanced using the Knight-Ruiz algorithm. The balanced cis matrixes were then normalized across samples being directly compared using median-of-ratios size factors conditioned on genomic distance. For trans interactions, because trans interactions are too sparse to quantify at higher matrix resolutions, each trans m x n contact matrix was assembled using Juicer by binning hg38 aligned, in situ Hi-C paired-end reads into uniform 1-Mb bins and then balanced using the Knight Ruiz algorithm with default parameters. Data was then quantile normalized across samples.
Assembly: hg38
Supplementary files format and content: The processed data files are the standard validPairs file format from the output of the software HiC-Pro. The data includes all paired reads that are valid.
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| Submission date |
Nov 23, 2022 |
| Last update date |
Dec 21, 2023 |
| Contact name |
Jennifer E Phillips-Cremins |
| E-mail(s) |
jcremins@seas.upenn.edu
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| Organization name |
University of Pennsylvania
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| Department |
Bioengineering
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| Street address |
415 Curie Blvd
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| City |
Philadelphia |
| State/province |
Pennsylvania |
| ZIP/Postal code |
19104 |
| Country |
USA |
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| Platform ID |
GPL18573 |
| Series (2) |
| GSE218675 |
Spatially coordinated heterochromatinization of long synaptic genes in fragile X syndrome [Hi-C] |
| GSE218680 |
Spatially coordinated heterochromatinization of long synaptic genes in fragile X syndrome |
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| Relations |
| BioSample |
SAMN31854609 |
| SRA |
SRX18368305 |
| Supplementary file |
Size |
Download |
File type/resource |
| GSM6754661_Proj025-FXSCYCLONE-JK-Exp9-FULCUM-NPC-HIC-111-1_merge_all_001_hg38.bwt2pairs.validPairs.txt.gz |
9.6 Gb |
(ftp)(http) |
TXT |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data provided as supplementary file |
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