|
| Status |
Public on Apr 10, 2014 |
| Title |
4P_0min_rep2 |
| Sample type |
RNA |
| |
|
| Channel 1 |
| Source name |
MW836a, mid-log
|
| Organism |
Saccharomyces cerevisiae |
| Characteristics |
genotype: HCM1 TOS4 YOX1 YHP1
time point: mid-log
|
| Growth protocol |
Cells were arrested in G1 phase with alpha-factor for 2 hours, released from the arrest, and samples collected at 15 minute intervals.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA was extracted using an acid-phenol prep as previously described (Schmitt et al, 1990; http://www.ncbi.nlm.nih.gov/pmc/articles/PMC330876/)
|
| Label |
Cy3
|
| Label protocol |
RNA was subjected to amplification and labeling using the Agilent Low Input Quick Amp Labeling Kit protocol (http://www.chem.agilent.com/library/usermanuals/Public/G4140-90050_GeneExpression_TwoColor_6.6.pdf) with minor modifications. Briefly, cRNA was amplified by in vitro transcription with amino-allyl UTP (3:2 ratio for amino-ally UTP: UTP) overnight at 37°C. Then, cRNA was purified using RNA Clean and Concentrator columns (Zymo Research) and labeled with Cy3 or Cy5 (GE healthcare) at room temperature for 90 minutes in the dark.
|
| |
|
| Channel 2 |
| Source name |
YBL289, alpha-factor arrest-release, 0 minutes
|
| Organism |
Saccharomyces cerevisiae |
| Characteristics |
genotype: hcm1-15A-3HA tos4-9A-3FLAG yox1-9A-3V5 yhp1-13A-13MYC
time point: 0 minutes
|
| Growth protocol |
Cells were arrested in G1 phase with alpha-factor for 2 hours, released from the arrest, and samples collected at 15 minute intervals.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA was extracted using an acid-phenol prep as previously described (Schmitt et al, 1990; http://www.ncbi.nlm.nih.gov/pmc/articles/PMC330876/)
|
| Label |
Cy5
|
| Label protocol |
RNA was subjected to amplification and labeling using the Agilent Low Input Quick Amp Labeling Kit protocol (http://www.chem.agilent.com/library/usermanuals/Public/G4140-90050_GeneExpression_TwoColor_6.6.pdf) with minor modifications. Briefly, cRNA was amplified by in vitro transcription with amino-allyl UTP (3:2 ratio for amino-ally UTP: UTP) overnight at 37°C. Then, cRNA was purified using RNA Clean and Concentrator columns (Zymo Research) and labeled with Cy3 or Cy5 (GE healthcare) at room temperature for 90 minutes in the dark.
|
| |
|
| |
| Hybridization protocol |
600ng of each cRNA was fragmented and hybridized as per manufacturers instructions (http://www.chem.agilent.com/library/usermanuals/Public/G4140-90050_GeneExpression_TwoColor_6.6.pdf).
|
| Scan protocol |
Slides were scanned on an Agilent G2505C scanner with ChipScan software version A.8.4.1.
|
| Description |
replicate 2 of 2
|
| Data processing |
Images were quantified using Agilent Feature Extraction Software (version 10.7.1.1) using default normalization settings.
|
| |
|
| Submission date |
Feb 18, 2014 |
| Last update date |
Apr 10, 2014 |
| Contact name |
Jennifer Benanti |
| E-mail(s) |
[email protected]
|
| Phone |
508-856-1773
|
| Organization name |
University of Massachusetts Medical School
|
| Department |
Molecular, Cell and Cancer Biology
|
| Street address |
364 Plantation St., LRB 525
|
| City |
Worcester |
| State/province |
MA |
| ZIP/Postal code |
01605 |
| Country |
USA |
| |
|
| Platform ID |
GPL16244 |
| Series (1) |
| GSE55121 |
Regulation of S-phase transcription factors by Cdk1 |
|
