|
| Status |
Public on Mar 18, 2013 |
| Title |
LB |
| Sample type |
RNA |
| |
|
| Source name |
MR-1 grown in rich LB medium
|
| Organism |
Shewanella oneidensis MR-1 |
| Characteristics |
treatment: grown in rich LB medium
media: Luria-Bertani broth (LB)
|
| Treatment protocol |
Bacteria were harvested at OD600nm~0.7 by centrifuging cultures for 10 minutes at 10,000 g at 4 C in RNAse free 50ml polypropylene tubes
|
| Growth protocol |
MR1 were grown at 30oC aerobically till mid exponential phase
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Bacterial pellets were collected by centrifuging cultures for 10 minutes at 10,000 g at 4 oC in RNAse free 50ml polypropylene tubes. Supernatant was immediately poured off and pellets were stored at -80 C. After thawing, RNA was extracted with RNeasy miniprep columns (Qiagen) with the optional on-column DNase treatment. RNA quality was confirmed with an Agilent Bioanalyzer; only samples with an RNA integrity number of around 8 or better were used. Ribosomal RNA was depleted with the MICROBExpress kit (Ambion), which uses magnetic beads coated with oligonucleotides that hybridize to ribosomal RNA.
|
| Label |
Alexa 555
|
| Label protocol |
First-strand cDNA was synthesized with random hexamer primers using SuperScript indirect cDNA labeling system (Invitrogen); the reaction buffer was supplemented with actinomycin D to inhibit second-strand synthesis. First-strand cDNA was labeled with Alexa 555. About 2 ug of labeled first-strand cDNA was hybridized to the array. For the genomic control, we used DNA from cells in stationary phase to minimize copy number variation across the chromosome. Genomic DNA was extracted using the DNeasy blood tissue kit (Qiagen) and labeled with Nimblegen's comparative genomic hybridization protocol. Briefly, genomic DNA was sonicated to 200-1000 bp and amplified using Klenow fragment and Cy3-labeled random nonamer primers.
|
| |
|
| Hybridization protocol |
Slides were hybridized using Nimblegen's standard protocol.
|
| Scan protocol |
Nimblegen slides were scanned on an Axon Gene Pix 4200A scanner with 100% gain
|
| Description |
cDNA
|
| Data processing |
Slides were analyzed with Nimblescan, with no local alignment and a border value of -1. Data from potentially cross-hybridizing probes (matching at 50 or more of 60 nucleotides to a second location in the genome) was removed.
|
| |
|
| Submission date |
Mar 15, 2013 |
| Last update date |
Mar 18, 2013 |
| Contact name |
Wenjun Shao |
| E-mail(s) |
[email protected]
|
| Organization name |
University of California
|
| Department |
Molecular and Cell Biology
|
| Lab |
Adam Arkin
|
| Street address |
2151 Berkeley Way
|
| City |
Berkeley |
| State/province |
CA |
| ZIP/Postal code |
94704-5230 |
| Country |
USA |
| |
|
| Platform ID |
GPL16797 |
| Series (2) |
| GSE45217 |
Transcript map of Shewanella oneidensis MR-1 |
| GSE58337 |
Conservation of transcription start sites within genes across a bacterial genus |
|
