|
| Status |
Public on Jul 06, 2010 |
| Title |
RNA2_v_gDNA |
| Sample type |
mixed |
| |
|
| Channel 1 |
| Source name |
Total_RNA_2
|
| Organism |
Kluyveromyces lactis |
| Characteristics |
strain: CLIB 209
|
| Treatment protocol |
50 ml of the culture were transferred to a 50 ml conical and spun down immediately. The isolated cell pellets were then placed in liquid nitrogen, stored at -80°C, and were later archived in RNAlater for future RNA extraction.
|
| Growth protocol |
Overnight cultures for each species were grown in 450ml of media at 220 RPM in a New Brunswick Scientific air-shaker at 30°C until reaching mid log-phase.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Qiagen Yeast RNA extraction
|
| Label |
Cy3
|
| Label protocol |
Indirect total RNA labeling
|
| |
|
| Channel 2 |
| Source name |
Genomic_DNA
|
| Organism |
Kluyveromyces lactis |
| Characteristics |
strain: CLIB 209
|
| Treatment protocol |
50 ml of the culture were transferred to a 50 ml conical and spun down immediately. The isolated cell pellets were then placed in liquid nitrogen, stored at -80°C, and were later archived in RNAlater for future RNA extraction.
|
| Growth protocol |
Overnight cultures for each species were grown in 450ml of media at 220 RPM in a New Brunswick Scientific air-shaker at 30°C until reaching mid log-phase.
|
| Extracted molecule |
genomic DNA |
| Extraction protocol |
Genomic DNA was isolated using Genomic-tip 500/G (Qiagen) using the provided protocol for yeast. DNA samples were sheared using Covaris sonicator to 500-1000 bp fragments, as verified using DNA 7500 and DNA 12000 kit for the Bioanalyzer 2100 (Agilent).
|
| Label |
Cy5
|
| Label protocol |
Genomic DNA samples were labeled with Cy5 using a modification of the protocol developed by Joe Derisi (UCSF) and Rosetta Inpharmatics (Kirkland, WA) that can be obtained at www.microarrays.org.
|
| |
|
| |
| Hybridization protocol |
Standard Agilent protocols for Agilent Custom Oligo Microarray 8x15K G4427A
|
| Scan protocol |
Arrays were scanned using an Agilent scanner and analyzed with Agilent’s feature extraction software version 10.5.1.1.
|
| Description |
252035410010_200812171227_S01_GE2-v5_10_Apr08_1_2
|
| Data processing |
For each probe, the median signal intensities were background subtracted for both channels and combined by taking the log2 of the Cy3 to Cy5 channel ratio. To estimate the absolute expression values for each gene, we took the median of the log2 ratios across all probes. The experiments were highly reproducible; most biological replicates correlated at R = 0.99 and replicates with R < 0.95 were removed.
|
| |
|
| Submission date |
Jun 08, 2010 |
| Last update date |
Jun 10, 2010 |
| Contact name |
Alex M Tsankov |
| E-mail(s) |
atsankov@mit.edu
|
| Phone |
6172532442
|
| Fax |
6172532442
|
| Organization name |
MIT
|
| Street address |
155 Bay State
|
| City |
Boston |
| State/province |
MA |
| ZIP/Postal code |
02215 |
| Country |
USA |
| |
|
| Platform ID |
GPL10499 |
| Series (2) |
| GSE21960 |
The role of nucleosome positioning in the evolution of gene regulation |
| GSE22198 |
Absolute expression level of Kluyveromyces lactis genes |
|
