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| Status |
Public on Aug 01, 2012 |
| Title |
HapMap MNase-seq GM18516 (Single-end) |
| Sample type |
SRA |
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| Source name |
lymphoblastoid cell line
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| Organism |
Homo sapiens |
| Characteristics |
cell type: lymphoblastoid cell line
population: YRI
ethnicity: Yoruba
country of origin: Nigeria
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| Biomaterial provider |
http://ccr.coriell.org/Sections/Search/Search.aspx?PgId=165&q=GM18516
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| Treatment protocol |
Cells were pelleted at 1000 rpm at 4C and washed in ice-cold PBS buffer. The cell pellet was resuspended in ice-cold NP-40 lysis buffer (10 mM Tris [pH 7.4], 10 mM NaCl, 3 mM MgCl2, 0.5% NP-40, 0.15 mM spermine, 0.5 mM spermidine) and incubated on ice for 5 minutes. The solution was centrifuged at 1000 rpm for 3 minutes at 4C, and then washed in a wash buffer (10 mM Tris [pH 7.4], 15 mM NaCl, 60 mM KCl, 0.15 mM spermine, 0.5 mM spermidine), and centrifuged again at 1000 rpm for 3 minutes at 4C. 1x10^8 cells were then resuspended in 10 ml of ice-cold MNase digestion buffer (10 mM Tris [pH 7.4], 15 mM NaCl, 60 mM KCl, 0.15 mM spermine, 0.5 mM spermidine, 1mM CaCl2).
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| Growth protocol |
Cell lines were cultured in RPMI 1640 media (supplemented with 2mM L-glutamine and 15% fetal bovine serum) and maintained at a density of between 2-5x10^5 viable cells/ml as per Coriell recommendations.
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| Extracted molecule |
genomic DNA |
| Extraction protocol |
1 ml aliquot of nuclei was digested with 7 μl of 50 U/μl MNase at 37°C for 12 minutes. Reaction was stopped by addition of EDTA, SDS and NaCl to an end concentration of 0.01M, 2% and 0.2M respectively. Reactions were digested with RNaseA (0.1mg) for 1hr at 42°C and further treated with ProteinaseK at 37°C for one hour. DNA was extracted using phenol-chloroform extraction and concentrated by ethanol precipitation. DNA was then run on a 3.3% Nusieve agarose gel at 75 V for 5 hours. 147bp fragments representing the mononucleosomes were excised from the gel and DNA was extracted from the gel by crushing the gel and soaking in soak buffer (300 mM Sodium Acetate, 1 mM EDTA, 0.1% SDS). The resulting DNA fragments in solution were then purified using Qiagen PCR purification kit. DNA fragments were prepared for paired-end sequencing using the standard Illumina protocol.
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| Library strategy |
MNase-Seq |
| Library source |
genomic |
| Library selection |
MNase |
| Instrument model |
Illumina HiSeq 2000 |
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| Description |
Single-end (SE) sequencing
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| Data processing |
Illumina CASAVA version 1.6 or 1.7 was used for base calling
Reads were mapped to the human genome with BWA version 0.5.8 or version 0.6.1-r104, using the commands 'bwa aln' and 'bwa sampe' or 'bwa samse' with default arguments.
SAM files were converted to BAM format, filtered for quality scores >= 10, sorted, and indexed using samtools version 0.1.13.
A python script was used to discarded reads if their fragment size fell outside the central 95% of the fragment size distribution (126-184 bp).
Midpoints of fragments were recorded using a python script. For single-end reads, the midpoint was assumed to be 75 bp from the 5’ end of the mapped read.
Genome_build: hg18
Supplementary_files_format_and_content: Wiggle files containing mapped MNase fragment midpoint counts for each base in the genome. Wiggle files are provided for each individual and for the combined data set. Wiggle files that only contain counts from only single-end (SE) reads or from paired-end reads from a defined fragment size range are also provided.
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| Submission date |
Apr 02, 2012 |
| Last update date |
May 15, 2019 |
| Contact name |
Graham McVicker |
| E-mail(s) |
gmcvicker@salk.edu
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| Organization name |
Salk Institute
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| Lab |
McVicker
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| Street address |
10010 N Torrey Pines Rd
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| City |
La Jolla |
| State/province |
CA |
| ZIP/Postal code |
92037 |
| Country |
USA |
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| Platform ID |
GPL11154 |
| Series (1) |
| GSE36979 |
Genome-wide maps of nucleosome occupancy in human lymphoblastoid cell lines |
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| Relations |
| SRA |
SRX134996 |
| BioSample |
SAMN00848618 |
| Named Annotation |
GSM907786_mnase_mids_NA18516_SE.wig.gz |
| Supplementary file |
Size |
Download |
File type/resource |
| GSM907786_mnase_mids_NA18516_SE.wig.gz |
267.4 Mb |
(ftp)(http) |
WIG |
SRA Run Selector |
| Raw data are available in SRA |
| Processed data provided as supplementary file |
| Processed data are available on Series record |
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