|
| Status |
Public on Sep 23, 2015 |
| Title |
77_MNase-3 nucleosome core particles (MNase) |
| Sample type |
SRA |
| |
|
| Source name |
S. cerevisiae
|
| Organism |
Saccharomyces cerevisiae |
| Characteristics |
strain name: JRY4012 (wild type)
genotype: MATa can1-100 his3-11,15 leu2-3,112 lys2-delta trp1-1 ura3-1
mnase or exoiii: MNase limit digest
|
| Treatment protocol |
Yeast nuclei were prepared from unfixed cells and then digested with a range of MNase concentrations at two different ExoIII concentrations. Mono-nucleosomal DNA was gel-purified (~150 bp). See paper for details.
In vitro reconstitutions were performed by mixing recombinant yeast histone octamers or native chicken erythrocyte core histones with purified plasmid pRS-ARG1-B DNA followed by salt dialysis. Reconstituted chromatin was digested to mono-nucleosomes using MNase and ExoIII. See paper for details.
Mouse nuclei were prepared from liver (female E13.5) and mildly digested with MNase only. The nuclei were lysed and soluble long chromatin was depleted of histone H1 using a cation exchange resin. Native chromatin and H1-depleted chromatin were digested with MNase and ExoIII. DNA from three digestion points (increasing MNase with fixed ExoIII) was purified. See paper for details.
|
| Growth protocol |
Yeast strains were grown to mid-log phase (OD600 about 0.8) at 30 degC in synthetic complete medium.
|
| Extracted molecule |
genomic DNA |
| Extraction protocol |
The DNA was repaired using the DNA repair kit from New England Biolabs. The repaired DNA was processed for paired-end sequencing according to the Illumina protocol.
MNase-seq and MNase-ExoIII-seq
|
| |
|
| Library strategy |
MNase-Seq |
| Library source |
genomic |
| Library selection |
MNase |
| Instrument model |
Illumina HiSeq 2000 |
| |
|
| Description |
Biological replicate 3
|
| Data processing |
Nucleosome sequences were aligned to the yeast genome (SacCer2) or the mouse genome (mm10) using Bowtie2 with default settings. Bowtie2 was also used to align the reconstituted nucleosome sequences to the plasmid sequence (supplied here).
This paper describes novel nucleosomal particles distinguished by their lengths. DNA fragment length distributions were calculated from the .sam files for all of the data sets and are supplied for the mouse data as text files.
Nucleosome occupancy maps were created using programs described by Cole et al. (2011) Nucleic Acids Res. 39, 9521-9535. Provided here are bigwig files for the yeast data and text files for the in vitro experiments with pRS-ARG1-B. Occupancy files were not calculated for the mouse nucleosomes, because the coverage is too low (~45m pairs of reads).
This paper describes novel nucleosomal particles distinguished by their lengths. DNA fragment length distributions were calculated from the .sam files for all of the data sets and are supplied for the mouse data as text files.
Genome_build: SacCer2 and mm10 for in vivo data. In vitro reconstitutions: the plasmid sequence (pRS-ARG1-B.fa) is provided here; the S. cerevisiae ARG1 locus (from -1283 to +2936 relative to the start codon) was inserted into pRS414. See paper for more details.
Supplementary_files_format_and_content: Yeast strains (in vivo data): bigwig files showing nucleosome occupancy (all reads included). Reconstituted nucleosomes on plasmid pRS-ARG1B: text files showing nucleosome occupancy at each plasmid coordinate (50-500 bp). Mouse data: length distributions (filenames contain "lengths"): text files with two columns: length (L) in bp, number of molecules of length L.
|
| |
|
| Submission date |
Sep 02, 2015 |
| Last update date |
May 15, 2019 |
| Contact name |
David Johannes Clark |
| E-mail(s) |
clarkda@mail.nih.gov
|
| Phone |
3014966966
|
| Organization name |
NICHD, NIH
|
| Department |
DDB
|
| Lab |
SCGE
|
| Street address |
6 Center Drive Bldg 6A Rm 2A02
|
| City |
Bethesda |
| State/province |
MD |
| ZIP/Postal code |
20892 |
| Country |
USA |
| |
|
| Platform ID |
GPL13821 |
| Series (1) |
| GSE65889 |
Novel nucleosomal particles containing core histones and linker DNA but no histone H1 |
|
| Relations |
| BioSample |
SAMN04027139 |
| SRA |
SRX1178196 |
