characteristics.text: ERC000011
characteristics.characteristic: ENA-CHECKLIST
characteristics.text: 2013-10-12T17:00:27Z
characteristics.characteristic: ENA-FIRST-PUBLIC
characteristics.text: 2018-03-08T16:49:40Z
characteristics.characteristic: ENA-LAST-UPDATE
characteristics.text: SAMEA2220305
characteristics.characteristic: External Id
characteristics.text: Department of Chemistry and Chemical Biology, Harvard University
characteristics.characteristic: INSDC center name
characteristics.text: 2013-10-12T17:00:27Z
characteristics.characteristic: INSDC first public
characteristics.text: 2018-03-08T16:49:40Z
characteristics.characteristic: INSDC last update
characteristics.text: public
characteristics.characteristic: INSDC status
characteristics.text: ERS358573
characteristics.characteristic: SRA accession
characteristics.text: E-MTAB-1950:iRFP_Nuc_3uM_40min
characteristics.characteristic: Submitter Id
characteristics.text: ArrayExpress
characteristics.ontologyTerms: http://www.ebi.ac.uk/efo/EFO_0009737
characteristics.characteristic: broker name
characteristics.text: baker's yeast
characteristics.characteristic: common name
characteristics.text: Protocols: The experimental protocol was essentially identical to what was previously described (Zhou & O'Shea, 2011). Briefly, 6 L of diploid (ASH79) was grown overnight in low fluorescence medium (-TRP -LEU) and then split into six 1L cultures. 40, 30, 20, 10, 5 and 0 min before the cultures reached an OD600 nm of 0.150 1-NM-PP1 was added to a final concentration of 3 uM. Cells were then crosslinked with 1% formaldehyde (final concentration) for 15 minutes at room temperature and quenched for 5 minutes with 125 mM glycine (final concentration). Cells were collected by centrifugation and immediately washed (10 mM Tris pH 7.5, 100 mM NaCl). The cell pellet was then dried and snap frozen in liquid nitrogen. The experimental protocol was essentially identical to what was previously described (Zhou & O'Shea, 2011). Briefly, 6 L of diploid (ASH79) was grown overnight in low fluorescence medium (-TRP -LEU) and then split into six 1L cultures. 40, 30, 20, 10, 5 and 0 min before the cultures reached an OD600 nm of 0.150 1-NM-PP1 was added to a final concentration of 3 uM. Cells were then crosslinked with 1% formaldehyde (final concentration) for 15 minutes at room temperature and quenched for 5 minutes with 125 mM glycine (final concentration). Cells were collected by centrifugation and immediately washed (10 mM Tris pH 7.5, 100 mM NaCl). The cell pellet was then dried and snap frozen in liquid nitrogen. The frozen cell pellet was thawed and 200 uL lysis buffer was added (50 mM HEPES, pH 7.5, 150 mM NaCl, 2 mM EDTA, 1% Triton X-100, 0.1% Sodium deoxycholate, complete protease inhibitor cocktail tablets (Roche cComplete Mini version 10, 1 tablet per 10 mL buffer)). The cells were then mechanically lysed with glass beads and bead beating. The cell pellets were then washed twice with MNase reaction buffer (10 mM Tris pH 7.5, 50 mM NaCl, 5 mM MgCl2, 1 mM CaCl2, 1 mM bME), resuspended in 750 uL MNase buffer and digested to primarily mononuclesomes with 0.75-1.5 U of MNase at 37 degrees C for 30 minutes. Subsequently samples were quenched by addition of 15 uL EDTA (500 mM). Debris was removed by centrifugation and 16 uL SDS (20%) and 30 uL proteinase K (10 mg/mL) were added to the supernatant. Crosslinks were then reversed and proteins digested by incubation at 65 degrees C overnight. The DNA was then purified twice by phenol/chloroform/isoamy alcohol extraction (25:24:1) and followed by precipitation in 1 mL ethanol at -20 degrees C. Precipitated DNA was resuspended in RNase A (0.2 mg/mL in TE buffer) and residual RNA removed by digesting for 1 hour at 37 degrees C. The samples were then purified using the PCR Purification Kit (Qiagen) and run on a 1.5% agarose gel for 60 minutes. The mononucleosome bands at ca. 150 bp were excised and purified on a Freeze 'N Squeeze DNA gel extraction column (BioRad). The DNA was then collected by ethanol precipitation. Sequencing libraries were prepared following the Illumina Multiplexed paired-end protocol. Adapters with a different index (0 min: ATCACG, 5 min: CGATGT, 10 min: TTAGGC, 20 min: TGACCA, 30 min: ACAGTG and 40 min: GCCAAT) for each timepoint were ligated onto the nucleosomal DNA fragments and the sequencing library amplified by 8 cycles of PCR.
characteristics.characteristic: description
characteristics.text: TPK1M164G TPK2M147G TPK3M165G msn4::TRP1/LEU2 MSN2-mCherry NHP6a-iRFP::kanMX
characteristics.characteristic: genotype
characteristics.text: Saccharomyces cerevisiae
characteristics.characteristic: organism
characteristics.text: Saccharomyces cerevisiae
characteristics.ontologyTerms: http://purl.obolibrary.org/obo/NCBITaxon_4932
characteristics.characteristic: organism
characteristics.text: diploid
characteristics.ontologyTerms: http://purl.obolibrary.org/obo/CL_0000415
characteristics.characteristic: ploidy
characteristics.text: Saccharomyces cerevisiae
characteristics.characteristic: scientific_name
characteristics.text: fresh specimen
characteristics.ontologyTerms: http://purl.obolibrary.org/obo/OBI_0000971
characteristics.characteristic: specimen with known storage state
characteristics.text: EY2807/ASH79
characteristics.characteristic: strain
characteristics.text: 40 minute
characteristics.ontologyTerms: http://purl.obolibrary.org/obo/UO_0000031
characteristics.characteristic: time
characteristics.text: iRFP_Nuc_3uM_40min
characteristics.characteristic: title