|
| Status |
Public on Aug 10, 2012 |
| Title |
Sinorhizobium meliloti podJ1 deletion mutant, biological replicate3 |
| Sample type |
RNA |
| |
|
| Source name |
podJ1 deletion mutant
|
| Organism |
Sinorhizobium meliloti |
| Characteristics |
genotype/variation: podJ1 deletion mutant
strain: JOE2553
|
| 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 |
For each biological replicate, a single bacterial colony was inoculated into 20 milliliters of LB medium, supplemented with streptomycin at 500 micrograms per microliter, and grown with shaking at 200 rpm at 30 degrees celsius in a 250 milliliter baffled flask to an OD-600 of 0.6 to 0.8
|
| 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 volume 101 pages 16636 to 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 |
The data were analyzed using Affymetrix software GCOS v1.4 with Affymetrix default analysis setting and global scaling as the normalization method. The mean target intensity of each array was arbitrarily set to 500. Affymetrix Data Mining Tool v3.1 was used for data mining as previously described by Barnett et al. 2004.
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| |
|
| Submission date |
Aug 10, 2011 |
| Last update date |
Aug 10, 2012 |
| 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 (1) |
| GSE31325 |
The Conserved Polarity Factor PodJ1 Impacts Multiple Cell Envelope-Associated Functions in Sinorhizobium meliloti |
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