|
| Status |
Public on Sep 01, 2012 |
| Title |
Salmonella_typhimurium_LT2_control_rep1 |
| Sample type |
RNA |
| |
|
| Source name |
Salmonella typhimurium LT2 control unexposed rep 1
|
| Organism |
Salmonella enterica subsp. enterica serovar Typhimurium str. LT2 |
| Characteristics |
genotype/variation: TA100
treatment: unexposed
|
| Growth protocol |
Salmonella typhimurium LT2 TA100 controls were grown in Davis Mingioli media supplemented with glucose (4 g/L), biotin (0.5 mg/L), and histidine (20 mg/L) for 24 h at 30°C shaking at 150 rpm.Davis BD, Mingioli ES. 1950. Mutants of Escherichia coli requiring methionine or vitamin B12. Journal of bacteriology 60: 17-28.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA was extracted using the RNeasy Mini kit (Qiagen Inc., Valencia, CA, USA) and according to the manufacturer’s instructions for RNA cleanup with an added enzymatic lysis step and mechanical disruption step added before cleanup. Enzymatic lysis and mechanical disruption was performed by resuspending the cell pellet in 100 uL TE buffer and 200 uL lysozyme (20 mg/mL), transferring the resuspended pellet to a MP Biomedicals (Solon, OH) Lysing Matrix B tube and vortexing for 1 min. The optional on column DNA digestion using the Qiagen RNase-Free DNase Kit was performed to remove DNA. RNA quality and concentration was determined by analysis with an Agilent 2100 bioanalyzer. The RNA was used to generate cDNA using the Superscript II Double-Stranded cDNA Synthesis Kit (Invitrogen, Carlsbad, CA) following the manufacturer’s directions. At each step the quality of the RNA and cDNA was analyzed using the Agilent 2100 Bioanalyzer and DNA 1000 Kit (Agilent).
|
| Label |
Cy3
|
| Label protocol |
The cDNA was labeled with the One-Color DNA Labeling Kit (Nimblegen, Madison, WI) and the concentration verified using the NanoDrop 1000 Spectrophotometer (Thermo Scientific, Waltham, MA).
|
| |
|
| Hybridization protocol |
Two micrograms of the labeled DNA was used for subsequent hybridization to Nimblegen Bacterial Gene Expression Arrays for Salmonella typhimurium LT2 X4. The hybridized arrays were washed using the Nimblegen Wash Buffer Kit.
|
| Scan protocol |
Washed arrays were scanned using the Agilent Microarray Scanner
|
| Description |
This sample is of Salmonella typhimurium LT2 strain TA100 control unexposed. It is the first of three biological replicates used in this experiment, each from separate cultures.
|
| Data processing |
Microarray images were analyzed using Nimblescan v2.5.26 software (Nimblegen) and ArrayStar v4.0.0 (DNAstar, Madison, WI) software. Expression data were log2 transformed and statistical significance was determined using a moderated t-test for binary transcriptome comparisons (i.e. control vs. experimental). The P-value was set at 0.05 and only those genes induced or repressed at least fivefold were considered in this paper. Differentially expressed transcripts were assigned to their functional classifications based on the Comprehensive Microbial Resource genome annotation from the J. Craig Venter Institute.
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| |
|
| Submission date |
Feb 13, 2012 |
| Last update date |
Sep 01, 2012 |
| Contact name |
Dawn E Hancock |
| E-mail(s) |
dawn.hancock@usace.army.mil
|
| Phone |
6016343711
|
| Organization name |
DEPARTMENT OF DEFENSE Army Corps of Engineers
|
| Department |
Engineer Research and Development Center
|
| Lab |
Environmental Lab
|
| Street address |
3909 Halls Ferry Road
|
| City |
Vicksburg |
| State/province |
Mississippi |
| ZIP/Postal code |
39180 |
| Country |
USA |
| |
|
| Platform ID |
GPL15224 |
| Series (1) |
| GSE35750 |
Expression analysis of Salmonella typhimurium LT2 TA100 upon exposure to C60 |
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