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Sample GSM230823 Query DataSets for GSM230823
Status Public on May 21, 2009
Title RH_1_rpd3_40 min
Sample type RNA
 
Channel 1
Source name BYrpd3 grown at 37C (time 0 reference)
Organism Saccharomyces cerevisiae
Characteristics Strain BY4741-rpd3:KanMX (MATa his3Δ1 leu2Δ0 met15Δ0 ura3Δ0 rpd3::KanMX)
Treatment protocol Stresses were added directly to the culture at the indicated concentrations. For heat shock experiments, cells were collected by rapid filtration at 25C and resuspended in fresh YPD heated to 37C. For reverse heat shock experiments, cells were grown 3 doublings in YPD at 37C, collected by brief centrifugation, and resuspended in fresh YPD at 25C
Growth protocol Unless otherwise noted, cells were grown 3 doublings in YPD or specified medium at 30C to OD600 ~0.5
Extracted molecule total RNA
Extraction protocol Hot-phenol RNA extraction as described in Gasch AP Methods in Enzymology 350: 393-414
Label Oyster 550 cyanine dye
Label protocol Indirect labeling using amino-allyl-dUTP and cyanine dyes from Flownamics (Madison, Wi) as described in Gasch AP Methods in Enzymology 350: 393-414
 
Channel 2
Source name BYrpd3 40 min after shift to 25C YPD
Organism Saccharomyces cerevisiae
Characteristics Strain BY4741-rpd3:KanMX (MATa his3Δ1 leu2Δ0 met15Δ0 ura3Δ0 rpd3::KanMX)
Treatment protocol Stresses were added directly to the culture at the indicated concentrations. For heat shock experiments, cells were collected by rapid filtration at 25C and resuspended in fresh YPD heated to 37C. For reverse heat shock experiments, cells were grown 3 doublings in YPD at 37C, collected by brief centrifugation, and resuspended in fresh YPD at 25C
Growth protocol Unless otherwise noted, cells were grown 3 doublings in YPD or specified medium at 30C to OD600 ~0.5
Extracted molecule total RNA
Extraction protocol Hot-phenol RNA extraction as described in Gasch AP Methods in Enzymology 350: 393-414
Label Oyster cyanine 650 dye
Label protocol Indirect labeling using amino-allyl-dUTP and cyanine dyes from Flownamics (Madison, Wi) as described in Gasch AP Methods in Enzymology 350: 393-414
 
 
Hybridization protocol As indicated in Gasch AP Methods in Enzymology 350: 393-414

To prepare the probe for hybridization to a ~22 x 22 mm microarray, combine 5/zl of each of the Cy-dUTP labeled samples or 10/zl of the combined aminoallyllabeled sample, 1/zl of 10 mg/ml poly(A) carrier (Sigma), 2.3/zl 20× SSC, and 0.36 tzl 10% SDS for a final volume of 14/zl (if the microarray is larger than 22 x 22 mm add to the 10 #1 of probe solution 2.6 #1 poly(A), 5.0 20x SSC, and 0.78 #1 10% SSD and adjust the final volume to 30 #1 with water). Denature the probe by heating at 94 ° for 3 min, then cool to room temperature by rapidly transferring to a microfuge and spinning for 10 sec (avoid adding bubbles during handling). Do not chill the probe on ice as salt will precipitate from the solution. Prepare the hybridization chamber (Coming): place the microarray into the chamber and add ~20/zl 3× SSC to each of the hybridization wells in the chamber and an additional -~30/zl of 3x SSC divided into isolated drops on the edges of the microarray. Insufficient volume will result in inadequate chamber hydration during hybridization, causing the probe to dry out under the coverslip. Select multiple microscope slide coverslips of the appropriate size (Coming) that appear free of dust, chips, and scratches, and prop them for easy access with a tweezers. Adding the probe to the microarray and sealing with the coverslip requires some experience and personal technique. It is advisable to practice the following procedure before hand. To the center of the microarray, deposit 12-15/zl of the denatured and cooled probe for 22 x 22 mm arrays, or 28-30 #1 probe for 22 x 44 mm microarrays. Bubbles can prevent proper hybridization, and therefore they should be avoided and removed when possible. Large bubbles can be popped with the comer of a coverslip, but great care should be taken not to touch the microarray surface. Quickly place the coverslip directly over the deposited probe and gently drop or place the coverslip over the array. The coverslip should not be moved so as to prevent scratching of the microarray surface; however, in unavoidable cases it can be gently positioned by sliding. Close the hybridization chamber and gently and quickly submerge the chamber in a 65 ° water bath. For optimal signal, hybridize the arrays for 7-16 hr. For consistent results, the hybridization conditions, including hybridization time, should be maintained for all microarrays.
Scan protocol Arrays were scanned on an Axon 4000B scanner
Description Comparison of rpd3 before and after reciprocal heat shock from 37C to 25C. Cells grown ~3 generations in YPD at 37C before centrifugation and resuspension in fresh YPD at 25C - replicate #2
Data processing Data were extracted using GenePix 6.0. Data were normalized by regional mean-centering as described by Lyne et al. 2003 BMC Genomics
 
Submission date Sep 20, 2007
Last update date Aug 14, 2011
Contact name Audrey P. Gasch
E-mail(s) [email protected]
Phone (608)265-0859
Fax (608)262-2976
Organization name University of Wisconsin Madison
Department Laboratory of Genetics
Lab Gasch Lab
Street address 425-g Henry Mall
City Madison
State/province WI
ZIP/Postal code 53706
Country USA
 
Platform ID GPL5915
Series (1)
GSE9108 The Histone Deacetylase Rpd3p Coordinates Transient Changes in Genomic Expression in Response to Acute Stress

Data table header descriptions
ID_REF
VALUE normalized, log2(sample/reference) expression ratios

Data table
ID_REF VALUE
YAR009C 0.4201
YAR010C 1.08
YBL005W-A 0.6001
YBL005W-B 0.3201
YBR012W-A 1.07
YBR012W-B 0.3701
YCL019W 0.1201
YCL020W 0.6001
YDR034C-C 0.6301
YDR034C-D 0.2201
YDR098C-A 1.17
YDR098C-B 0.1001
YDR170W-A 1.04
YDR210W-A 0.7001
YDR210W-B 0.3601
YDR261C-C 1.05
YDR261C-D 0.5701
YDR261W-A 0.6301
YDR261W-B 0.2601
YDR316W-A 1.07

Total number of rows: 6308

Table truncated, full table size 91 Kbytes.




Supplementary file Size Download File type/resource
GSM230823.gpr.gz 692.0 Kb (ftp)(http) GPR
Processed data included within Sample table

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