|
| Status |
Public on Sep 05, 2018 |
| Title |
Sinorhizobium meliloti CL150 overexpressing RpoE1, biological rep 2 |
| Sample type |
RNA |
| |
|
| Source name |
wild type strain CL150 (ecfR1 pstC mutations corrected) carrying pF1087 (RpoE1)
|
| Organism |
Sinorhizobium meliloti |
| Characteristics |
genotype/variation: overexpressing RpoE1
strain: CL150 pF1087
|
| Treatment protocol |
Stop solution (5% buffer equilibrated phenol in ethanol) was added to bacterial cultures, at one tenth the culture volume, before centrifugation at 4 degrees celsius. Bacterial cell pellets were frozen in liquid nitrogen and stored at -80 degrees celsius until RNA extraction.
|
| Growth protocol |
Bacteria were cultured in M9 medium with 0.4% glycerol as a carbon source, and supplemented with 50 ug/ml spectinomycin and 500 ug/ml streptomycin. When cultures reached an OD-600 of 0.5, melibiose was added to 0.4% for 30 minutes.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA was extracted using a Qiagen RNeasy kit with modifications as described by Barnett et al. 2004 (Proc Natl Acad Sci USA 101:16636-16641).
|
| Label |
biotin
|
| Label protocol |
cDNA was fragmented and biotinylated as described by Barnett et al. 2004.
|
| |
|
| Hybridization protocol |
Four micrograms of biotinylated cDNA was hybridized to each chip at 48 degrees celsius for 16 hours as described by Barnett et al. 2004. GeneChips were washed and stained in the Affymetrix Fluidics Station 400
|
| Scan protocol |
Chips were scanned at the Stanford PAN facility using an Affymetrix scanner 3000 7G
|
| Data processing |
Affymetrix CEL files were analyzed using R and BIOCONDUCTOR software packages (Gentleman et al. 2004, Genome Biol 5:R80). Probe summarization and normalization were carried out with the AFFY package and the RMA algorithm (Irizarry et al. 2003, Biostatistics 4: 249-264).
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| |
|
| Submission date |
Jul 05, 2018 |
| Last update date |
Sep 07, 2018 |
| Contact name |
Melanie J Barnett |
| Organization name |
Stanford University
|
| Department |
Biology
|
| Lab |
Sharon R. Long
|
| Street address |
371 Jane Stanford Way
|
| City |
Stanford |
| State/province |
CA |
| ZIP/Postal code |
94305 |
| Country |
USA |
| |
|
| Platform ID |
GPL9757 |
| Series (2) |
| GSE116665 |
Overexpression of Sinorhizobium meliloti ECF sigma factors. |
| GSE116680 |
Most Sinorhizobium meliloti ECF-sigma factors perform accessory functions |
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