|
| Status |
Public on Aug 18, 2009 |
| Title |
totRNA_UV_2 |
| Sample type |
RNA |
| |
|
| Source name |
total RNA extracted 20 min after induction of the SOS response by UV irradiation (50 J/m2)
|
| Organism |
Escherichia coli str. K-12 substr. MG1655 |
| Characteristics |
strain: K-12 MG1655
cell culture: mid-log phase
|
| Treatment protocol |
50 ml culture of Escherichia coli K-12 MG1655 was irradiated by UV 50 J/m2 OD600 = 0.5. The culture was further harvested by centrifugation 20 minutes after exposure to UV. (Samples 6-10)
|
| Growth protocol |
Escherichia coli K-12 MG1655 single colony from overnight cultures were diluted 1:500 in 100 ml K-medium (1xM9, 1.2% glucose, 1.25% casamino acids (dCAA), 1 mM MgSO4, 0.1 mM CaCl2) and grown at 37°C with aeration by rotary shaking until OD600=0.5.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
1 ml trizol was added pr 106 cells and incubated at room temperature for 5 minutes. 0.2 ml chloroform was added pr ml Trizol and the sample was mixed for 15 seconds before 15 minutes centrifutgation (12000 g 4°C). The aqueous phase was slowly added 1:1(vol) to 70% EtOH and further loaded to a RNeasy column (QIAGEN), washed and DNase treated according to RNeasy protocol (QIAGEN).
|
| Label |
Biotin
|
| Label protocol |
Fragmented cDNA was terminal labelled with Biotin according to the Affymetrix 'Prokaryotic sample and array processing' protocol version 701029 Rev. 4.
|
| |
|
| Hybridization protocol |
Fragmented and labelled cDNA were hybridized to arrays overnight at 45°C according to the Affymetrix 'Prokaryotic sample and array processing' protocol version 701029 Rev. 4.
|
| Scan protocol |
The arrays were scanned using GeneChip Skanner 3000 7G with settings appropriate to the array type.
|
| Description |
single colony and culture parallell to totRNA_MOCK_2 (the culture was split in two, half induced by UV irradiation the other half as reference was not induced)
|
| Data processing |
Details on the analysis methods applied is found in the connected paper: Custom design and analysis of high-density oligonucleotide bacterial tiling microarrays. Thomassen GO, Rowe AD, Lagesen K, Lindvall JM, Rognes T. PLoS One. 2009 Jun 17;4(6):e5943. PMID: 19536279
Results of analysis of the reference dataset (the 5 MOCK E. coli arrays,) and the treated dataset (the 5 UV treated E. coli arrays) are provided in the "MOCKvsUVresults.tar" supplementary file available on the Series record.
|
| |
|
| Submission date |
Dec 04, 2008 |
| Last update date |
Sep 24, 2010 |
| Contact name |
Gard Thomassen |
| E-mail(s) |
[email protected]
|
| Phone |
+4793674926
|
| URL |
http://cmbn.no/rognes
|
| Organization name |
Centre for Molecular Biology and Neuroscience (CMBN) and Institute of Medical Microbiology
|
| Department |
Molecular microbiology
|
| Lab |
Bioinformatics
|
| Street address |
Rikshospitalet University Hospital
|
| City |
Oslo |
| State/province |
Oslo |
| ZIP/Postal code |
0027 |
| Country |
Norway |
| |
|
| Platform ID |
GPL7714 |
| Series (1) |
| GSE13829 |
UV treated Escherichia coli reveals a high number of differentially expressed regions and many novel transcripts |
|
