|
| Status |
Public on Mar 22, 2011 |
| Title |
hupB_6h_Replicate 2 |
| Sample type |
mixed |
| |
|
| Channel 1 |
| Source name |
hupB, 6h
|
| Organism |
Salmonella enterica subsp. enterica serovar Typhimurium |
| Characteristics |
strain: SL1344
genotype: delta-hupB
time: 6 hours
|
| Growth protocol |
To prepare cells for RNA extraction, an overnight culture of the relevant strain was grown under antibiotic selection where appropriate, and used to inoculate 1:100 25 ml of fresh antibiotic free LB in a 250 ml flask incubated with shaking at 250 rpm in a New Brunswick Innova 3100 waterbath at 37C. Samples were removed for RNA extraction at 1 h, 4 h and 6 h after inoculation.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA isolation was performed using the Promega SV Total RNA Purification kit (Cat. No.: Z3100). For details, see the protocol at http://www.ifr.ac.uk/safety/Microarrays/protocols.html.
|
| Label |
Cy5
|
| Label protocol |
Labelling protocol at:
http://www.ifr.ac.uk/safety/Microarrays/protocols.html.
|
| |
|
| Channel 2 |
| Source name |
Reference genomic DNA
|
| Organism |
Salmonella enterica subsp. enterica serovar Typhimurium |
| Characteristics |
strain: SL1344
genotype: wild type
|
| Growth protocol |
To prepare genomic DNA, cells were grown in LB under aeration until stationary phase.
|
| Extracted molecule |
genomic DNA |
| Extraction protocol |
Genomic DNA was isolated using the Qiagen 'Genomic DNA' Kit (Cat. No.: 19060 for the buffer kit; 10243 for the columns).
|
| Label |
Cy3
|
| Label protocol |
Labelling protocol at:
http://www.ifr.ac.uk/safety/Microarrays/protocols.html.
|
| |
|
| |
| Hybridization protocol |
Slides blocked with DCE according to protocol at PMID 11266573. For hybridisation and washing protocols, see http://www.ifr.ac.uk/safety/Microarrays/protocols.html.
|
| Scan protocol |
Axon GenePix4000A, 10um resolution scan, and see http://www.ifr.ac.uk/safety/Microarrays/protocols.html.
|
| Description |
hupB_6h: Biological Replicate 2 of 2.
035746B_hupB6.2
|
| Data processing |
Data centering was performed by bringing the median ratio (Cy5/Cy3) for each of the 16 blocks to one using the following equation: (Ti) = (Cy5i/Cy3i) - c, where T is the centered ratio, i is the gene index, Cy5 and Cy3 are the red and green intensities and c is the 50th percentile of all Cy5/Cy3 ratios. Flagged features are omitted from the data centering process. Because we used a microarray based on S. Typhimurium strain LT2 to analyse the transcriptional response of a different strain (S. Typhimurium strain SL1344), we removed all the data corresponding to genes present in LT2, but absent from SL1344.
|
| |
|
| Submission date |
Jul 09, 2010 |
| Last update date |
Mar 23, 2011 |
| Contact name |
Sacha Lucchini |
| E-mail(s) |
sacha.lucchini@bbsrc.ac.uk
|
| Organization name |
Institute of Food Research
|
| Department |
Molecular Microbiology
|
| Street address |
Colney Lane
|
| City |
Norwich |
| ZIP/Postal code |
NR4 7UA |
| Country |
United Kingdom |
| |
|
| Platform ID |
GPL9091 |
| Series (1) |
| GSE22860 |
The nucleoid-associated protein HU controls three regulons that coordinate virulence and general physiology in Salmonella enterica serovar Typhimurium |
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