|
| Status |
Public on Aug 07, 2013 |
| Title |
ura3 (2+) t=-60 |
| Sample type |
RNA |
| |
|
| Source name |
ura3, 60 minutes before glucose
|
| Organism |
Halobacterium salinarum NRC-1 |
| Characteristics |
strain: ∆ura3 (WT)
timepoint: -60
rna amount (ng): 100
|
| Treatment protocol |
Addition of glucose to 5% at t=0
|
| Growth protocol |
Gene expression was assayed in a previously constructed strain containing an in-frame deletion of VNG1451C (∆ura3∆trmB; Schmid et al., 2009) and its isogenic parent strain (∆ura3). Cells for gene expression were grown in a Complete Defined Medium (CDM), containing 19 amino acids (modified from Schmid et al., 2009) supplemented with 50 mM uracil to complement the ura3 deletion. Cultures were grown at 225 r.p.m. shaking at 42°C under low ambient light.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Cells were immediately pelleted by centrifugation and snap frozen in liquid nitrogen. RNA was prepared using the Absolutely-RNA miniprep kit (Stratagene/Agilent) according to the manufacturer’s instructions. RNA quality was assessed using the Agilent 2100 BioAnalyzer using the RNA-Nano Chip and the Prokaryotic Nano RNA protocol, and samples were verified to be free of DNA contamination by 25 cycles of PCR amplification on at least 200 ng RNA prior to cDNA conversion in RT-qPCR.
|
| Label |
none
|
| Label protocol |
No labeling
|
| |
|
| Hybridization protocol |
Performed by Nanostring (see Geiss, et at. 2008)
|
| Scan protocol |
Performed by Nanostring (see Geiss, et at. 2008)
|
| Description |
Cartridge 4
|
| Data processing |
Genes were normalized by the sum of the positive spike in controls. Genes were then normalized by the geometric mean of the three reference genes (srp19, mrp, vng1670c). Resulting numbers were further normalized by the average sum of all counts per strain (ura3, trmB) in the 100ng samples. Microsoft Excel was used for this normalization.
|
| |
|
| Submission date |
May 22, 2013 |
| Last update date |
Aug 07, 2013 |
| Contact name |
Horia Todor |
| E-mail(s) |
[email protected]
|
| Organization name |
Duke University
|
| Department |
Biology
|
| Lab |
Schmid
|
| Street address |
125 Science Drive
|
| City |
Durham |
| State/province |
North Carolina |
| ZIP/Postal code |
27708 |
| Country |
USA |
| |
|
| Platform ID |
GPL17188 |
| Series (1) |
|
