|
| Status |
Public on Nov 02, 2007 |
| Title |
Transcriptional profiling of Loc1 knockout replicate #1 |
| Sample type |
RNA |
| |
|
| Channel 1 |
| Source name |
Loc1 knockout cells grown in rich media at 30C
|
| Organism |
Saccharomyces cerevisiae |
| Characteristics |
Strain: PSY3262
ura3 leu2-3,112 his3-11 trp1-1 ade2-1 ho can1-100 LOC1::KAN
|
| Growth protocol |
50mL cultures were grown in YPD to early log phase (OD600 0.2-0.4) at 30oC, centrifuged at room temeperature and flash-frozen using liquid nitrogen.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
mRNA was extracted from cell pellets using the hot phenol method and precipitated overnight in ethanol
|
| Label |
Cy5
|
| Label protocol |
mRNA was reverse-transcribed using a poly-T probe, and then labeled with Cy5 using Klenow and random primers.
|
| |
|
| Channel 2 |
| Source name |
Wild-type cells grown in rich media at 30C
|
| Organism |
Saccharomyces cerevisiae |
| Characteristics |
Strain: PSY3259
ura3 leu2-3,112 his3-11 trp1-1 ade2-1 ho can1-100
|
| Growth protocol |
50mL cultures were grown in YPD to early log phase (OD600 0.2-0.4) at 30oC, centrifuged at room temeperature and flash-frozen using liquid nitrogen.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
mRNA was extracted from cell pellets using the hot phenol method and precipitated overnight in ethanol
|
| Label |
Cy3
|
| Label protocol |
mRNA was reverse-transcribed using a poly-T probe, and then labeled with Cy3 using Klenow and random primers.
|
| |
|
| |
| Hybridization protocol |
Sample was mixed with DIG Easy Hybe, salmon sperm DNA, and yeast tRNA, and incubated at 65oC for 2 minutes. Sample was then hybridized to microarray and incubated at 37oC for 16 hours. Arrays were washed three times for ten minutes each with 1xSSC, 0.1%SDS at 50oC. Arrays were then briefly rinsed with 1X SSC at 50oC and air-dried in a centrifuge.
|
| Scan protocol |
The Cy3 and Cy5 intensities at each spot were determined using a GenePix 4000B scanner.
|
| Description |
Replicate #1 of transcriptional profiling of Loc1- cells.
|
| Data processing |
The ratio of 635/532 was determined at each pixel using Genepix. Value is equal to the median of the ratios for a given spot.
|
| |
|
| Submission date |
Aug 13, 2007 |
| Last update date |
Aug 14, 2011 |
| Contact name |
Suzanne Komili |
| Organization name |
Harvard University
|
| Street address |
44 Binney St.
|
| City |
Boston |
| State/province |
MA |
| ZIP/Postal code |
02115 |
| Country |
USA |
| |
|
| Platform ID |
GPL5741 |
| Series (1) |
| GSE8765 |
Transcriptional profiling of Loc1 knockout |
|
