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Sample GSM832850 Query DataSets for GSM832850
Status Public on Jan 01, 2012
Title sgs1_BrdU_HU_60_min
Sample type genomic
 
Channel 1
Source name BrdU-IP from sgs1 deletion mutant cells in HU, 60 min
Organism Saccharomyces cerevisiae
Characteristics strain: W303
antibody: BrdU
vendor/catalog: BDBioscience 555627
genotype/variation: sgs1 deletion mutant cells
time point: 60 min
Growth protocol G1-synchronised cells were released into S phase in the presence of Hydroxyurea (200mM) and BrdU (400µg/ml) for the indicated time.
Extracted molecule genomic DNA
Extraction protocol For immunoprecipitation of BrdU-labelled DNA, 1.5x109 cells were lysed for 5 x 2 minutes in NIB buffer (17 % glycerol, 50 mM MOPS buffer, 150mM potassium acetate, 2 mM magnesium chloride, 500 µM spermidine and 150 µM spermine, pH7.2) with Zirconium beads on a Vibrax (VXR basic, Ika) at 4C. DNA was isolated using Qiagen Genomic DNA extraction kit and sonicated to reach an average DNA size of 300-600 bp. For each immunoprecipitation, 10 µl of mouse anti-BrdU IgG1 (BDBioscience 555627) was prebound to Dynabeads M-280 Sheep anti-mouse IgG (Invitrogen 112.02) and added to the denatured purified DNA. For immunoprecipitation of BrdU-labelled DNA, 1.5x109 cells were lysed for 5 x 2 minutes in NIB buffer (17 % glycerol, 50 mM MOPS buffer, 150mM potassium acetate, 2 mM magnesium chloride, 500 µM spermidine and 150 µM spermine, pH7.2) with Zirconium beads on a Vibrax (VXR basic, Ika) at 4C. DNA was isolated using Qiagen Genomic DNA extraction kit and sonicated to reach an average DNA size of 300-600 bp. For each immunoprecipitation, 10 µl of mouse anti-BrdU IgG1 (BDBioscience 555627) was prebound to Dynabeads M-280 Sheep anti-mouse IgG (Invitrogen 112.02) and added to the denatured purified DNA. For immunoprecipitation of BrdU-labelled DNA, 1.5x109 cells were lysed for 5 x 2 minutes in NIB buffer (17 % glycerol, 50 mM MOPS buffer, 150mM potassium acetate, 2 mM magnesium chloride, 500 µM spermidine and 150 µM spermine, pH7.2) with Zirconium beads on a Vibrax (VXR basic, Ika) at 4C. DNA was isolated using Qiagen Genomic DNA extraction kit and sonicated to reach an average DNA size of 300-600 bp. For each immunoprecipitation, 10 µl of mouse anti-BrdU IgG1 (BDBioscience 555627) was prebound to Dynabeads M-280 Sheep anti-mouse IgG (Invitrogen 112.02) and added to the denatured purified DNA. For immunoprecipitation of BrdU-labelled DNA, 1.5.109 cells were lysed for 5 x 2 min in NIB buffer (17% glycerol, 50 mM MOPS buffer, 150 mM potassium acetate, 2 mM magnesium chloride, 500 µM spermidine and 150 µM spermine, pH 7.2) with zirconium beads on a vibrax (VXR basic, IKA) at 4°C. DNA was isolated using Qiagen Genomic DNA extraction kit and sonicated to reach an average DNA size of 300-600 bp. For each immunoprecipitation, 10µl of mouse anti-BrdU IgG1 (BD Biosciences) was prebound to Dynabeads M-280 Sheep anti-mouse IgG (Invitrogen) and added to the denaturated purified DNA.
Label biotin
Label protocol 7 µg of amplified DNA was fragmented and labelled using the GeneChip WT Double-Stranded DNA Terminal Labelling kit (Affymetrix, PN 900812) following manufacturer's recommendations. 7 µg of amplified DNA was fragmented and labelled using the GeneChip WT Double-Stranded DNA Terminal Labelling kit (Affymetrix, PN 900812) following manufacturer recommendations.
 
Channel 2
Source name Genomic DNA from Saccharomyces cerevisiae
Organism Saccharomyces cerevisiae
Characteristics strain: W303
fraction: input
Growth protocol G1-synchronised cells were released into S phase in the presence of Hydroxyurea (200mM) and BrdU (400µg/ml) for the indicated time.
Extracted molecule genomic DNA
Extraction protocol For immunoprecipitation of BrdU-labelled DNA, 1.5x109 cells were lysed for 5 x 2 minutes in NIB buffer (17 % glycerol, 50 mM MOPS buffer, 150mM potassium acetate, 2 mM magnesium chloride, 500 µM spermidine and 150 µM spermine, pH7.2) with Zirconium beads on a Vibrax (VXR basic, Ika) at 4C. DNA was isolated using Qiagen Genomic DNA extraction kit and sonicated to reach an average DNA size of 300-600 bp. For each immunoprecipitation, 10 µl of mouse anti-BrdU IgG1 (BDBioscience 555627) was prebound to Dynabeads M-280 Sheep anti-mouse IgG (Invitrogen 112.02) and added to the denatured purified DNA. For immunoprecipitation of BrdU-labelled DNA, 1.5x109 cells were lysed for 5 x 2 minutes in NIB buffer (17 % glycerol, 50 mM MOPS buffer, 150mM potassium acetate, 2 mM magnesium chloride, 500 µM spermidine and 150 µM spermine, pH7.2) with Zirconium beads on a Vibrax (VXR basic, Ika) at 4C. DNA was isolated using Qiagen Genomic DNA extraction kit and sonicated to reach an average DNA size of 300-600 bp. For each immunoprecipitation, 10 µl of mouse anti-BrdU IgG1 (BDBioscience 555627) was prebound to Dynabeads M-280 Sheep anti-mouse IgG (Invitrogen 112.02) and added to the denatured purified DNA. For immunoprecipitation of BrdU-labelled DNA, 1.5x109 cells were lysed for 5 x 2 minutes in NIB buffer (17 % glycerol, 50 mM MOPS buffer, 150mM potassium acetate, 2 mM magnesium chloride, 500 µM spermidine and 150 µM spermine, pH7.2) with Zirconium beads on a Vibrax (VXR basic, Ika) at 4C. DNA was isolated using Qiagen Genomic DNA extraction kit and sonicated to reach an average DNA size of 300-600 bp. For each immunoprecipitation, 10 µl of mouse anti-BrdU IgG1 (BDBioscience 555627) was prebound to Dynabeads M-280 Sheep anti-mouse IgG (Invitrogen 112.02) and added to the denatured purified DNA. For immunoprecipitation of BrdU-labelled DNA, 1.5.109 cells were lysed for 5 x 2 min in NIB buffer (17% glycerol, 50 mM MOPS buffer, 150 mM potassium acetate, 2 mM magnesium chloride, 500 µM spermidine and 150 µM spermine, pH 7.2) with zirconium beads on a vibrax (VXR basic, IKA) at 4°C. DNA was isolated using Qiagen Genomic DNA extraction kit and sonicated to reach an average DNA size of 300-600 bp. For each immunoprecipitation, 10µl of mouse anti-BrdU IgG1 (BD Biosciences) was prebound to Dynabeads M-280 Sheep anti-mouse IgG (Invitrogen) and added to the denaturated purified DNA.
Label biotin
Label protocol 7 µg of amplified DNA was fragmented and labelled using the GeneChip WT Double-Stranded DNA Terminal Labelling kit (Affymetrix, PN 900812) following manufacturer's recommendations. 7 µg of amplified DNA was fragmented and labelled using the GeneChip WT Double-Stranded DNA Terminal Labelling kit (Affymetrix, PN 900812) following manufacturer recommendations.
 
 
Hybridization protocol DNA was hybridized to Affymetrix Genechip S. cerevisiae Tiling 1.0R Array (5 bpresolution, Affymetrix PN 900645) using the GeneChip Hybridization, Wash, and Stain Kit (Affymetrix, PN 900720).
Scan protocol Tiling arrays were scanned with the GeneChip scanner 3000 7G
Data processing results file descriptions: signal bar files generated for each strains contains log2 (IP/Input) for signal intensity.
 
Submission date Nov 14, 2011
Last update date Jan 01, 2012
Contact name Philippe Pasero
E-mail(s) ppasero@igh.cnrs.fr
Organization name Institute of Human Genetics - CNRS
Lab Pasero
Street address 141 rue de la Cardonille
City Montpellier
ZIP/Postal code 34396
Country France
 
Platform ID GPL7250
Series (1)
GSE33686 dNTP pools determine fork progression and origin usage under replication stress

Supplementary file Size Download File type/resource
GSM832850_JP012_WCEA.CEL.gz 32.1 Mb (ftp)(http) CEL
GSM832850_LC029_WCEA.CEL.gz 34.8 Mb (ftp)(http) CEL
GSM832850_sgs1delta_JP080_IP7.CEL.gz 27.7 Mb (ftp)(http) CEL
GSM832850_sgs1delta_JP080_IP7_signal.bar.gz 15.6 Mb (ftp)(http) BAR
Processed data provided as supplementary file

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