|
| Status |
Public on Mar 02, 2023 |
| Title |
WT_42_2 vs delta-sigB_42_2 |
| Sample type |
RNA |
| |
|
| Channel 1 |
| Source name |
WT_42_2
|
| Organism |
Bacillus cereus ATCC 14579 |
| Characteristics |
sample type: Bacillus cereus ATCC14579 WT grown in BHI at 30 degrees C up to OD600nm of 0.4-0.5 followed by 20 minutes of growth at 42 degrees C, shaken at 160 rpm
|
| Treatment protocol |
The mid log cultures (OD600nm ~0.4-0.5) are exposed, remaining in the same media, for 20 minutes at 30 degrees C (reference temperature) or 20 minutes at 42 degrees C (heat shock)
|
| Growth protocol |
Cells are precultured in BHI at 30 degrees C, shaking at 160 rpm, untill an OD600nm reached between 0.4-0.5 before exposure
|
| Extracted molecule |
total RNA |
| Extraction protocol |
RNA was extracted as described by van Schaik et al., 2004
|
| Label |
Cy5
|
| Label protocol |
Complementary DNA was synthesized and labeled using 15 μg of total RNA, and the CyScribe Post-Labeling kit (Amersham Biosciences Europe GmbH) according to the supplier's instructions.
|
| |
|
| Channel 2 |
| Source name |
delta-sigB_42_2
|
| Organism |
Bacillus cereus ATCC 14579 |
| Characteristics |
sample type: Bacillus cereus ATCC14579 delta-sigB grown in BHI at 30 degrees C up to OD600nm of 0.4-0.5 followed by 20 minutes of growth at 42 degrees C, shaken at 160 rpm
|
| Treatment protocol |
The mid log cultures (OD600nm ~0.4-0.5) are exposed, remaining in the same media, for 20 minutes at 30 degrees C (reference temperature) or 20 minutes at 42 degrees C (heat shock)
|
| Growth protocol |
Cells are precultured in BHI at 30 degrees C, shaking at 160 rpm, untill an OD600nm reached between 0.4-0.5 before exposure
|
| Extracted molecule |
total RNA |
| Extraction protocol |
RNA was extracted as described by van Schaik et al., 2004
|
| Label |
Cy3
|
| Label protocol |
Complementary DNA was synthesized and labeled using 15 μg of total RNA, and the CyScribe Post-Labeling kit (Amersham Biosciences Europe GmbH) according to the supplier's instructions.
|
| |
|
| |
| Hybridization protocol |
The target was denatured by boiling for 1 min and immediate cooling on ice prior to hybridization. A total of 150ng of both samples was mixed and applied with the Agilent hybridization buffer to the microarray, according to Agilent manual. Hybridization was performed at 60 °C. The slide was washed according to the Agilent microarray wash protocol.
|
| Scan protocol |
8 x 15K Microarrays were scanned with the Agilent DNA Microarray Scanner, Model G2505C, using the extended dynamic range scan mode and 10 micron scan resolution according to the Agilent protocol.
|
| Description |
WT 42 degrees C vs delta-sigB 42 degrees C, biological duplo, 2 of 2
|
| Data processing |
Images were analyzed and processed with the Agilent Feature Extraction software (version 10.7.3.1). The obtained extracted data files were analyzed using the limma software package [39] in R (R Core Team, 2014). For Agilent-based arrays, global loess normalization was used when the bulk of genes is not differentially expressed. The analysis was based on a 2-color experimental design using a linear modeling approach by lmfit and empirical Bayes statistics [41].
Log 2 of Lowess normalized ratio (wildtype vs mutants)(30 degrees C over 42 degrees C)
Lowess normalized ratio (wildtype vs mutants)(30 degrees C over 42 degrees C)
|
| |
|
| Submission date |
Sep 22, 2022 |
| Last update date |
Mar 02, 2023 |
| Contact name |
Jeroen Steven Koomen |
| E-mail(s) |
Jeroen.Koomen@wur.nl
|
| Organization name |
Wageningen University
|
| Lab |
Food Microbiology
|
| Street address |
Bornse Weilanden 9
|
| City |
Wageningen |
| ZIP/Postal code |
6708 WG |
| Country |
Netherlands |
| |
|
| Platform ID |
GPL9493 |
| Series (1) |
| GSE213958 |
SigB modulates expression of novel SigB regulon members via Bc1009 in non-stressed and heat-stressed cells revealing its alternative roles in Bacillus cereus |
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