|
| Status |
Public on Oct 30, 2012 |
| Title |
Hi-C Progeria (HGPS) Fibroblasts p19 |
| Sample type |
SRA |
| |
|
| Source name |
patient forearm skin biopsy, Hi-C
|
| Organism |
Homo sapiens |
| Characteristics |
disease status: Hutchinson-Gilford progeria syndrome
tissue: forearm skin biopsy
cell type: fibroblasts
gender: male
cell line: HGADFN167
passage: 19
|
| Treatment protocol |
20 million cells were crosslinked in 1% formaldehyde for each Hi-C experiment.
|
| Growth protocol |
Primary human dermal fibroblasts were cultured in MEM (Invitrogen/GIBCO) supplemented with 15% fetal bovine serum (FBS) (Invitrogen) and 2 mM L-glutamine.
|
| Extracted molecule |
genomic DNA |
| Extraction protocol |
Hi-C was carried out essentially as described previously (Lieberman-Aiden et al., 2009 [PMID 19815776]). Cells were lysed and chromatin was digested with HindIII, then digested ends were filled in with biotinylated dCTP and ligated for 4 hours at 16 C. After reversing the formaldehyde crosslinks by incubation at 65 C with Proteinase K overnight, unligated biotinylated ends were removed by incubation with 15 U T4 DNA polymerase.
The ligation products were fragmented by Covaris sonication to an average size of 200 bp, and then the ideal size for Illumina sequencing (100-300 bp) was selected by Ampure fractionation (0.9x Ampure beads to remove the DNA fragments >300 bp followed by 1.3x Ampure beads to isolate the remaining DNA fragments > 100 bp). The DNA ends were prepared for Illumina sequencing adapter ligation by repairing the DNA ends and adding an 'A' to each end. The biotinylated junctions were then pulled down using MyOne streptavidin beads at a ratio of 1 uL beads per ng of biotinylated DNA. Illumina paired end adapters were ligated onto the DNA ends and then the fragments were PCR amplified. Samples were sequenced on an Illumina GAII instrument using the Paired End 75 bp module.
|
| |
|
| Library strategy |
OTHER |
| Library source |
genomic |
| Library selection |
other |
| Instrument model |
Illumina Genome Analyzer II |
| |
|
| Description |
HGPS-p19
HGPS patient.
|
| Data processing |
Library strategy: Hi-C
Reads were mapped to the hg18 genome using Bowtie2 using the "very-sensitive" settings in an iterative procedure, starting with the first 25 bp of the sequence and iterating to the full 75 bp length as previously described (Imakaev et al., Nature Methods, 2012 [22941365]).
Aligned reads were assigned to restriction fragments and filtered to discard duplicate read pairs (PCR over-amplification products) and molecules for which both ends map to the same restriction fragment. Restriction fragments shorter than 100 bp or longer than 100kb, as well as those with the top 0.5% of read counts, were removed (validPair file).
Reads were assigned to genomic bins of 200 kb, according to the center of their corresponding restriction fragment. The binned interaction maps were then corrected for systematic biases by equalizing the total coverage (1D sum across the matrix) of every bin in the genome using 50 iterations of a normalization procedure previously described (Imakaev et al., Nature Methods, 2012) (IntMatrix file). These matrices were then smoothed with a 1 Mb bin size and 200 kb step size for publication figures.
Genome_build: hg18
Supplementary_files_format_and_content: validPair files: Comment lines describing the inputs to the file are all preceeded by a '#' character. The 3 columns of data (tab delimited) are: 1) The first restriction fragment of the interacting pair (restriction fragment ID number|coordinates); 2) The second restriction fragment of the interacting pair; 3) The total number of unique paired end reads that map to this combination of restriction fragments.
Supplementary_files_format_and_content: IntMatrix files: Contains a matrix of normalized interaction frequencies for every pair of 200 kb bins in the genome. Bin names and genomic coordinates are listed in row and column headers, and then iteratively corrected interaction frequencies are listed in each pair of row and column.
|
| |
|
| Submission date |
Oct 22, 2012 |
| Last update date |
May 15, 2019 |
| Contact name |
Rachel Patton McCord |
| E-mail(s) |
Rachel.McCord@umassmed.edu
|
| Phone |
508-856-4377
|
| Organization name |
University of Massachusetts Medical School
|
| Department |
Program in Gene Function and Expression
|
| Lab |
Job Dekker Lab
|
| Street address |
364 Plantation St. LRB 570M
|
| City |
Worcester |
| State/province |
MA |
| ZIP/Postal code |
01605 |
| Country |
USA |
| |
|
| Platform ID |
GPL9115 |
| Series (2) |
| GSE41763 |
Correlated alterations in genome organization, histone methylation, and DNA-lamina interactions in Hutchinson-Gilford progeria syndrome (Hi-C) |
| GSE41764 |
Correlated alterations in genome organization, histone methylation, and DNA-lamina interactions in Hutchinson-Gilford progeria syndrome |
|
| Relations |
| SRA |
SRX200035 |
| BioSample |
SAMN01774248 |
