|
| Status |
Public on Jan 02, 2008 |
| Title |
Saccharopolyspora RNA at 36h biological rep.A |
| Sample type |
RNA |
| |
|
| Source name |
RNA extracted from Saccharopolyspora erythraea after 36h of growth
|
| Organism |
Saccharopolyspora erythraea |
| Characteristics |
strain: NRRL 2338
|
| Treatment protocol |
No treatment.
|
| Growth protocol |
A dense culture of Sac. erythraea grown in oat meal agar was inoculated into SCM liquid medium (20 g soytone, 15 g soluble starch, 10.5 g morpholinepropanesulfonic acid, 1.5 g yeast extract and 0.1 g CaCl2 per liter of distilled H2O) and grown for 48 h at 33ºC at 200 rpm. The culture was then stored as 1-ml frozen aliquots, which were individually used to inoculate three independent 500 ml buffled flasks each containing 100 ml SCM medium. After 12 h at 33ºC and 200 rpm, 1 ml samples, from each flask, were withdrawn, separating the mycelium from the broth by centrifugation.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
For each time point, RNA was extracted from mycelium pellets deriving fron 1 ml culture samples using the GeneElute™ total RNA Purification Kit (SIGMA), recovering it in 50 µl of Elution Solution.
|
| Label |
biotin
|
| Label protocol |
The RNAs were processed following the Affymetrix Protocol Target Labeling for Prokaryotic GeneChip® Antisense Arrays (Affymetrix Procaryotic gene Expression Manual). The protocol consists in cDNA synthesis by reverse transcription (starting with 10 µg RNA), followed by cDNA fragmentation with DNase I and labelling with Terminal Deoxynucleotidyl Transferase.
|
| |
|
| Hybridization protocol |
The labelled cDNAs were then hybridised for 16h at 50°C on individual GeneChips
|
| Scan protocol |
GeneChips were washed and stained with streptavidin-conjugated phycoerythrin by using the Fluidic Station FS450 (Affymetrix) following the ProkGE-WS2v3_450 Protocol. Fluorescent images of the microarrays were acquired using a GeneChip Scanner 3000 (Affymetrix).
|
| Description |
Gene expression data from Saccharopolyspora erythraea after 36h of growth
|
| Data processing |
Raw data quality was assessed considering MAS5.0 control parameters after a global scaling at a target intensity of 100. Quality and control parameters as well as boxplot of raw intensities indicated the overall high quality of the data set and the absence of any outlying sample. Probe level data was converted to expression values using both Robust Multi-array Average procedure and MAS5.0 algorithms. In the former case, PM values have been background adjusted, normalized using invariant set normalization, and log transformed. In the latter case, intensity levels have been normalized using the Global Scaling option to target value (i.e., TGT=100).
|
| |
|
| Submission date |
Oct 24, 2007 |
| Last update date |
Aug 14, 2011 |
| Contact name |
Giorgio Corti |
| E-mail(s) |
[email protected]
|
| Organization name |
National Research Council
|
| Department |
Institute of Biomedical Technologies
|
| Street address |
via Fratelli Cervi 93
|
| City |
Segrate - Milan |
| ZIP/Postal code |
20090 |
| Country |
Italy |
| |
|
| Platform ID |
GPL6051 |
| Series (1) |
| GSE9422 |
Complete Gene Expression Profiling of Saccharopolyspora erythraea NRRL 2338 |
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