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| Status |
Public on Aug 28, 2011 |
| Title |
Hop1 rec8delta mutant, 4h meiosis, biological repeat 1 |
| Sample type |
genomic |
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| Channel 1 |
| Source name |
Hop1 ChIP with anti -Hop1 antibody in rec8delta mutant
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| Organism |
Saccharomyces cerevisiae |
| Characteristics |
genetic background: SK1
genotype: FKY3683: Mat a/alpha, REC114-myc13::KanMX6/=, rec8D::KanMX/=
bait protein: Hop1
antibody: rabbit, anti-Hop1
meiotic timepoint: 4 hours
genotype: wild type
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| Growth protocol |
Meiotic cultures of diploid S. cerevisiae SK1 strain were set up as follows: A saturated YPD culture was used to inoculate 300 ml liquid SPS medium (.5% yeast extract, 1% Bacto peptone, 1% potassium acetate, .17% Yeast nitrgen base w/o aa, 0.05 M potassium biphtalate, pH 5.5) 1:400 and grown in a 3-liter baffled flask at 200 rpm at 30°C for 14 hrs. Cells were harvested, washed once, resuspended at an OD660 of 1.1 in sporulation medium (1% potassium acetate pH 7.0, supplemented with amino acids and .001% PPG) and shaken at 200 rpm at 30°C for 4 hrs.
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| Extracted molecule |
genomic DNA |
| Extraction protocol |
Cells were fixed in 1% formaldehyde for 30min. 1X10^9 cells were disrupted by MULTI-BEADS SHOCKER (MB400U, YASUI KIKAI, Osaka). Chromosomal DNA was shared into the average size of 500-1000bp by sonication. Chromatin IP was performed as described in Katou et al. (Methods Enzym., 409,389-410, 2006).
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| Label |
Biotin-11-ddATP
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| Label protocol |
Two rounds of amplification, DNA fragmentation, and end-labeling, were performed according to the Affymetrix ChIP protocol.
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| Channel 2 |
| Source name |
WCE 4h meiosis
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| Organism |
Saccharomyces cerevisiae |
| Characteristics |
genotype: rad50S mutation
genetic background: SK1
bait protein: none
antibody: none
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| Growth protocol |
Meiotic cultures of diploid S. cerevisiae SK1 strain were set up as follows: A saturated YPD culture was used to inoculate 300 ml liquid SPS medium (.5% yeast extract, 1% Bacto peptone, 1% potassium acetate, .17% Yeast nitrgen base w/o aa, 0.05 M potassium biphtalate, pH 5.5) 1:400 and grown in a 3-liter baffled flask at 200 rpm at 30°C for 14 hrs. Cells were harvested, washed once, resuspended at an OD660 of 1.1 in sporulation medium (1% potassium acetate pH 7.0, supplemented with amino acids and .001% PPG) and shaken at 200 rpm at 30°C for 4 hrs.
|
| Extracted molecule |
genomic DNA |
| Extraction protocol |
Cells were fixed in 1% formaldehyde for 30min. 1X10^9 cells were disrupted by MULTI-BEADS SHOCKER (MB400U, YASUI KIKAI, Osaka). Chromosomal DNA was shared into the average size of 500-1000bp by sonication. Isolation of genomic DNA was performed as described in Katou et al. (Methods Enzym., 409,389-410, 2006).
|
| Label |
Biotin-11-ddATP
|
| Label protocol |
Two rounds of amplification, DNA fragmentation, and end-labeling, were performed according to the Affymetrix ChIP protocol.
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| Hybridization protocol |
Hybridization to Affymetrix S. cerevisiae Tiling 1.0R Arrays was carried out at 42C for 16 hours. The following washing and staining steps were done with an Affymetrix 450 station. All reagents used are described in the Affymetrix ChIP Assay Protocol.
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| Scan protocol |
Each array was scanned by Genechip Scanner 3000 (Affymetrix, CA) at default settings
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| Data processing |
Primary data analysis was performed with GeneChip Operating Software (GCOS, Affymetrix). Resulting Cel files were analyzed by the Tiling Analysis Software (TAS, v 1.1.02, Affymetrix, two sample comparison analysis (IP versus WCE), without normalization and results in linear scale were exported as RPT text file) in the format specified by Affymetrix.
Text files are in tab separated format provided by Affymetrix Tiling Analysis Software. The first column represents the chromosomal position, the second column "fold enrichment" over the control experiment (WCE, 4 hours, if not specified otherwise). The columns are separated by 25 headers specifying the identity of the chromosome; sequence #8-#23 contain chromosome1 to chromosome16, sequence #24 mitochondrial DNA, sequence #25 the yeast 2mikron plasmid.
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| Submission date |
Jun 09, 2011 |
| Last update date |
Aug 28, 2011 |
| Contact name |
Franz Klein |
| E-mail(s) |
franz.klein@univie.ac.at
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| Phone |
++431427756220
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| Organization name |
University of Vienna, MFPL
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| Department |
Chromosome Biology
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| Street address |
Dr. Bohrg. 9
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| City |
Vienna |
| ZIP/Postal code |
A-1030 |
| Country |
Austria |
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| Platform ID |
GPL7250 |
| Series (1) |
| GSE29860 |
Spo11-accessory proteins link DNA double-strand break sites to the chromosome axis in early meiotic recombination |
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| Supplementary file |
Size |
Download |
File type/resource |
| GSM739691_190310_3683_rec8del_Hop1_2.CEL.gz |
31.0 Mb |
(ftp)(http) |
CEL |
| GSM739691_190310_3683_rec8del_Hop1_2.txt.gz |
15.4 Mb |
(ftp)(http) |
TXT |
| GSM739691_WCE_4h_meiosis_copy23.CEL.gz |
28.5 Mb |
(ftp)(http) |
CEL |
| Processed data provided as supplementary file |
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