|
| Status |
Public on Dec 31, 2011 |
| Title |
Sigma70 LP-1 |
| Sample type |
RNA |
| |
|
| Channel 1 |
| Source name |
Sigma70 LP
|
| Organism |
Escherichia coli |
| Characteristics |
transformation: pSigma70-LP
treatment: 42 C
genotype: 285c rpoD mutation
|
| Treatment protocol |
Control samples (285c/empty vector) were harvested via filtration and resuspended in RNALater once they had reached OD = 0.5. Experimental samples were harvested after 1 hour of heat shock at 42 C in an analogous procedure.
|
| Growth protocol |
All strains were grown in M9 minimal media supplemented with 25 ug/ml streptomycin and 0.52 mM arginine. Samples were grown to OD = 0.5 @ 30 C; experimental samples were then transferred to a shaking water bath @ 42 C.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
The Zymo Fungal/Bacterial RNA extraction kit was used to extract RNA from all samples. In-column DNaseI treatment was used to eliminate genomic DNA carryover as described by the manufacturer.
|
| Label |
Cy3
|
| Label protocol |
The Fairplay III Kit (Stratagene) and Cy3/Cy5 dyes (Amersham) were used to generate labeled cDNA for microarray hybridization
|
| |
|
| Channel 2 |
| Source name |
285c/control vector at 30 C 1
|
| Organism |
Escherichia coli |
| Characteristics |
transformation: empty vector
treatment: 30 C
genotype: 285c rpoD mutation
|
| Treatment protocol |
Control samples (285c/empty vector) were harvested via filtration and resuspended in RNALater once they had reached OD = 0.5. Experimental samples were harvested after 1 hour of heat shock at 42 C in an analogous procedure.
|
| Growth protocol |
All strains were grown in M9 minimal media supplemented with 25 ug/ml streptomycin and 0.52 mM arginine. Samples were grown to OD = 0.5 @ 30 C; experimental samples were then transferred to a shaking water bath @ 42 C.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
The Zymo Fungal/Bacterial RNA extraction kit was used to extract RNA from all samples. In-column DNaseI treatment was used to eliminate genomic DNA carryover as described by the manufacturer.
|
| Label |
Cy5
|
| Label protocol |
The Fairplay III Kit (Stratagene) and Cy3/Cy5 dyes (Amersham) were used to generate labeled cDNA for microarray hybridization
|
| |
|
| |
| Hybridization protocol |
Hybridization occurred as described by the Agilent two-color prokaryotic microarray protocol (18 hours total time).
|
| Scan protocol |
Arrays were washed according to the Agilent gene expression microarray protocol and then subsequently scanned using an Axon Genepix 4200A microarray scanner.
|
| Description |
RNA, heat shocked for 1 hour
|
| Data processing |
Data were normalized using LOWESS (MIDAS) and analyzed for statistical significance using the Rank Product method in the MultiExperimentViewer
|
| |
|
| Submission date |
Apr 01, 2011 |
| Last update date |
Dec 31, 2011 |
| Contact name |
James Winkler |
| E-mail(s) |
jdwinkler@tamu.edu
|
| Organization name |
Texas A&M University
|
| Lab |
Kao Lab
|
| Street address |
3122 TAMU
|
| City |
College Station |
| State/province |
TX |
| ZIP/Postal code |
77843 |
| Country |
USA |
| |
|
| Platform ID |
GPL13359 |
| Series (1) |
| GSE28320 |
Characterization of Lactobacillus Sigma Factors using a Temperature Sensitive E. coli Sigma70 Mutant |
|
