|
| Status |
Public on Aug 01, 2014 |
| Title |
Scc2-4 25°C Mnase-Seq |
| Sample type |
SRA |
| |
|
| Source name |
Mono-nucleosomal DNA
|
| Organism |
Saccharomyces cerevisiae |
| Characteristics |
strain: Y4251
genotype: MATa, scc2-4, Scc1-Pk9::TRP1, Sth1-9myc::URA3
|
| Treatment protocol |
Cells were cross-linked with 1% Formaldehide for 30 minutes at room temperature. The reaction was stopped with 0.125 M Glycine for 10 minutes.
|
| Growth protocol |
Cells were grown in YP + 2% Glucose to an OD600 of 0,4. Then the cells were incubated for two more hours at the indicated temperature.
|
| Extracted molecule |
genomic DNA |
| Extraction protocol |
Preparation of mono-nucleosomal DNA was performed as described (Lantermann et al. 2009) followed by high throughput sequencing on Illumina GAIIx and HiSeq 2500. Briefly, cells were grown under the specified conditions and then fixed with 1% formaldehyde. Spheroblasts were obtained and the chromatin digested with 30u/ml of Mnase (Fermentas) at 37°C for 20 minutes. The samples were then treated with RNase and Proteinase K to isolate the undigested, mono-nucleosomal DNA. The DNA was then size selected and purified in a 1.5% agarose gel and precipitated with ethanol.
Sequencing libraries were prepared according to the manufacturer's protocol (15005180 Rev. A and 15026486 Rev. B).
|
| |
|
| Library strategy |
MNase-Seq |
| Library source |
genomic |
| Library selection |
MNase |
| Instrument model |
Illumina Genome Analyzer IIx |
| |
|
| Description |
Genomic DNA obtained after digestion with MNase to isolate mono-nucleosomal DNA
|
| Data processing |
Read alignment was performed using the Burrows-Wheeler Aligner, bwa (version 0.5.9-r16). If required, prior to alignment, the sacCer2 release of the budding yeast genome was modified to reflect genome alterations that were introduced for the experiment.
Discordantly mapped read-pairs were removed, leaving only those that were paired, had a maximum of 2 mismatches in any given read and an insert size between 140 and 170 bp.
Genome-wide wig files were generated by treating each pair of reads as a single fragment.
Genome_build: Saccharomyces cerevisiae genome (version sacCer2)
Supplementary_files_format_and_content: Genome-wide wig files were generated at 1bp resolution by treating each pair of reads as a single fragment, including the inferred insert portion. A total library size of 25 million read pairs was used to normalise the coverage for cross-sample comparison.
|
| |
|
| Submission date |
Apr 23, 2014 |
| Last update date |
May 15, 2019 |
| Contact name |
Lidia Lopez-Serra |
| E-mail(s) |
lidia.lopez-serra@cancer.org.uk
|
| Organization name |
Canser Research UK
|
| Department |
Chromosome segregation
|
| Street address |
44 Lincoln's Inn Fields
|
| City |
London |
| ZIP/Postal code |
WC2A 3LY |
| Country |
United Kingdom |
| |
|
| Platform ID |
GPL13272 |
| Series (2) |
| GSE56992 |
The Scc2NIPBL/Scc4MAU2 complex acts in sister chromatid cohesion and transcriptional regulation by maintaining nucleosome-free regions [Mnase-seq] |
| GSE56994 |
The Scc2NIPBL/Scc4MAU2 complex acts in sister chromatid cohesion and transcriptional regulation by maintaining nucleosome-free regions. |
|
| Relations |
| BioSample |
SAMN02732224 |
| SRA |
SRX523893 |
