|
| Status |
Public on May 21, 2018 |
| Title |
SPT6-TET-H3_dox_2 |
| Sample type |
genomic |
| |
|
| Channel 1 |
| Source name |
SPT6-TET-H3_dox
|
| Organism |
Saccharomyces cerevisiae |
| Characteristics |
strain background: BY4741
genotype/variation: SPT6-TET
epitope tag or untagged genotype: SPT6-TET promoter
treated with: Sulfometuron methyl + doxycycline
antibody used: H3 (Abcam: 1791)
molecule subtype: ChIP DNA
|
| Treatment protocol |
The cells were grown in SC and induced for Gcn4, by treating cells grown in SC with with 0.65 µM of sulfometuron methyl (SM; Chemservice, cat# N-13254) for 20 minutes and processed for ChIP analysis. The kin28as cells were treated with 1NA-PP1 before processing for cross-linking and chromatin preparation. For depleting Spt6 or Spt16, SPT16-TET and SPT6-TET cells were cultured in the presence of 10µg/ml of doxycycline.
|
| Growth protocol |
Chromatin immunoprecipitation experiments were performed as described previously (Govind et al., 2012). Briefly, 100 ml of cells (A600 = 0.6) were cross-linked with 1% formaldehyde for 15 minutes at ambient temperature and quenched with glycine. Chromatin was isolated and fragmented by sonication (Branson450) to an average size of 300-400 base pairs. The soluble fraction of chromatin was used for ChIP using the appropriate antibodies.
|
| Extracted molecule |
genomic DNA |
| Extraction protocol |
The ChIP and Input DNA was extracted by Phenol:Chloform method after reverse-crosslinking at 65 degree C, and after proteinase K treatment. DNA was fragmented with DNase prior to labeling
|
| Label |
Alexa555
|
| Label protocol |
The manufacturer's recommended protocol was followed.
|
| |
|
| Channel 2 |
| Source name |
SPT6-TET-H3_dox_input
|
| Organism |
Saccharomyces cerevisiae |
| Characteristics |
strain background: BY4741
genotype/variation: SPT6-TET
name: SPT6-TET
molecule subtype: Input genomic DNA
|
| Treatment protocol |
The cells were grown in SC and induced for Gcn4, by treating cells grown in SC with with 0.65 µM of sulfometuron methyl (SM; Chemservice, cat# N-13254) for 20 minutes and processed for ChIP analysis. The kin28as cells were treated with 1NA-PP1 before processing for cross-linking and chromatin preparation. For depleting Spt6 or Spt16, SPT16-TET and SPT6-TET cells were cultured in the presence of 10µg/ml of doxycycline.
|
| Growth protocol |
Chromatin immunoprecipitation experiments were performed as described previously (Govind et al., 2012). Briefly, 100 ml of cells (A600 = 0.6) were cross-linked with 1% formaldehyde for 15 minutes at ambient temperature and quenched with glycine. Chromatin was isolated and fragmented by sonication (Branson450) to an average size of 300-400 base pairs. The soluble fraction of chromatin was used for ChIP using the appropriate antibodies.
|
| Extracted molecule |
genomic DNA |
| Extraction protocol |
The ChIP and Input DNA was extracted by Phenol:Chloform method after reverse-crosslinking at 65 degree C, and after proteinase K treatment. DNA was fragmented with DNase prior to labeling
|
| Label |
Alexa647
|
| Label protocol |
The manufacturer's recommended protocol was followed.
|
| |
|
| |
| Hybridization protocol |
The samples were labeled using BioPrime Array CGH Genomic Labeling Module kit and hybridized according to the Manufacturer's recommended protocol.
|
| Scan protocol |
Agilent Technologies Scanner G2505B US45102973
|
| Description |
Biological Replicate-2
|
| Data processing |
Data processed on R. Median normalization carried out.
|
| |
|
| Submission date |
Jun 08, 2015 |
| Last update date |
May 21, 2018 |
| Contact name |
Chhabi Govind |
| E-mail(s) |
govind@oakland.edu
|
| Organization name |
Oakland University
|
| Department |
Biological Sciences
|
| Street address |
333 Science and Engineering Building
|
| City |
Rochester |
| State/province |
mi |
| ZIP/Postal code |
48085 |
| Country |
USA |
| |
|
| Platform ID |
GPL10930 |
| Series (1) |
| GSE69642 |
Spatial regulation of transcription and histone occupancy by histone chaperones FACT and Spt6 |
|
