|
| Status |
Public on Mar 01, 2021 |
| Title |
IAA_IN_4h_YKR1894 |
| Sample type |
SRA |
| |
|
| Source name |
Cells
|
| Organism |
Saccharomyces cerevisiae |
| Characteristics |
strain: YKR1894
genome used for alignment: sacCer3
|
| Treatment protocol |
Site-specific DSBs were introduced inducing the HO endonuclease or the AsiSI restriction enzyme via the galactose promoter, adding galactose to the cultures to a final concentration of 2%. Cells were crosslinked at the indicated time points with 1% formaldehyde for 16 minutes at room temperature and the reaction quenched with glycine (final concentration 400 mM). Cells were harvested by centrifugation (3500xg at 4°C), washed with cold PBS and the pellet flash frozen in liquid nitrogen.
|
| Growth protocol |
Cells were grown in YP-media containing 2% raffinose at 30°C. Exponentially growing cells (OD600 0.5-0.7) were synchronized in M phase with 5 μg/ml nocodazole for 2-3 hours. Cell cycle arrest was assessed via microscopy and DNA content measured by flow cytometry.
|
| Extracted molecule |
genomic DNA |
| Extraction protocol |
Cells were mechanically lysed and chromatin was sheared by sonication to fragments of 200-500 bp. 40% of the chromatin extract was used for immunoprecipitation, the purified complexes were treated with proteinase K and crosslinks reversed. DNA was purified by phenol-chloroform extraction and ethanol precipitation.
Strand-specific ChIP-seq libraries were prepared starting from 1-3 ng of DNA using Accel-NGS® 1S Plus Library Kit following manufacturer instructions and 10-12 cycles were used for library amplification. Clean-up steps were performed with SPRIselect beads.
|
| |
|
| Library strategy |
ChIP-Seq |
| Library source |
genomic |
| Library selection |
ChIP |
| Instrument model |
Illumina NextSeq 500 |
| |
|
| Data processing |
Sequecing reads were mapped to the S. cerevisiae genome using Bowtie2 (version 2.2.9) with default parameters
Multiple mapping reads were filtered with SAMtools.
Genome_build: LYZE00000000.1, sacCer3
Supplementary_files_format_and_content: All files were generated with R. Bed files contain aligned reads, bedGraph files represent read coverage
|
| |
|
| Submission date |
Jan 05, 2021 |
| Last update date |
Mar 01, 2021 |
| Contact name |
Martina Peritore |
| E-mail(s) |
peritore@biochem.mpg.de
|
| Phone |
+49 8985783049
|
| Organization name |
MPIB
|
| Lab |
Pfander
|
| Street address |
Am Klopferspitz 18
|
| City |
Planegg-Martinsried |
| State/province |
Bayern |
| ZIP/Postal code |
82152 |
| Country |
Germany |
| |
|
| Platform ID |
GPL19756 |
| Series (1) |
| GSE149807 |
Strand-specific ChIP-seq at DNA breaks distinguishes single versus doubled stranded DNA binding and refutes single stranded nucleosomes |
|
| Relations |
| BioSample |
SAMN17218457 |
| SRA |
SRX9786326 |
