|
| Status |
Public on Mar 01, 2013 |
| Title |
Treatment B Population 5 Colony c replica 2 |
| Sample type |
RNA |
| |
|
| Channel 1 |
| Source name |
Pure culture in LB
|
| Organism |
Escherichia coli |
| Characteristics |
strain: MC1000
oxygen regime: daily shift between shaking and static (no shaking) conditions
population: 5
morphology: c
culture type: evolved
|
| Growth protocol |
Growth curves of all selected evolved colony types and the ancestor were obtained by growing them under their evolved environmental conditions in LB medium until Log-phase. For this, optical density was recorded at 600nm at a value of 0.6 to 0.7.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Cells of each evolved colony type were collected by centrifugation at 13,000 × g for 1 min at 4°C and were immediately used for total RNA isolation with NucleoSpin RNA II isolation kit (Macherey – Nagel, Biokè, Leiden, the Netherlands).
|
| Label |
Cy5
|
| Label protocol |
High quality RNA samples were reverse-transcribed into cDNA with the Superscript III reverse transcriptase kit (Invitrogen, Carlsbad, USA) and labeled with Dylight Amine-Reactive Dyes Cy3 and Cy5 (Thermo Scientific, Rockford, USA).
|
| |
|
| Channel 2 |
| Source name |
Pure culture in LB
|
| Organism |
Escherichia coli |
| Characteristics |
strain: MC1000
culture type: Ancestor
|
| Growth protocol |
Growth curves of all selected evolved colony types and the ancestor were obtained by growing them under their evolved environmental conditions in LB medium until Log-phase. For this, optical density was recorded at 600nm at a value of 0.6 to 0.7.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Cells of each evolved colony type were collected by centrifugation at 13,000 × g for 1 min at 4°C and were immediately used for total RNA isolation with NucleoSpin RNA II isolation kit (Macherey – Nagel, Biokè, Leiden, the Netherlands).
|
| Label |
Cy3
|
| Label protocol |
High quality RNA samples were reverse-transcribed into cDNA with the Superscript III reverse transcriptase kit (Invitrogen, Carlsbad, USA) and labeled with Dylight Amine-Reactive Dyes Cy3 and Cy5 (Thermo Scientific, Rockford, USA).
|
| |
|
| |
| Hybridization protocol |
Hybridization in a dye-swap design was performed using the Agilent Gene Expression Hybridization Kit (Agilent technologies CA, USA) at 65°C for 17 h to genome-wide multi-strain E. coli (8x15K) microarrays of 60-mer oligonucleotides (Agilent Technologies, CA, USA). Two microarrays in the slides were used per evolved colony type (two biological replicas). Each replica was labeled with a different dye (cy3 or cy5) and hybridazed to the ancestors labeled with the oposite dye.
|
| Scan protocol |
Scanning was performed using GenePix autoloader 4200AL confocal laser scanner (Molecular devices Ltd.) according to the provider's protocol. http://www.moleculardevices.com
|
| Data processing |
Statistical analysis was performed using R Limma package (Linear Models for Microarrays Data) (Smyth 2004), where the two microarrays per sample were normalized using lowess normalization method. Multiple-gene probes results were merged and normalized using the MA table conversion tool available on the MOLGEN Bioinformatics Server (http://server.molgenrug.nl/ - Molecular genetics, university of Groningen, the Netherlands).
|
| |
|
| Submission date |
Feb 25, 2013 |
| Last update date |
Mar 01, 2013 |
| Contact name |
Pilar E Puentes-Téllez |
| E-mail(s) |
[email protected]
|
| Organization name |
University of Groningen
|
| Department |
Microbial Ecology
|
| Street address |
Nijenborgh 7
|
| City |
Groningen |
| ZIP/Postal code |
9747AG |
| Country |
Netherlands |
| |
|
| Platform ID |
GPL13360 |
| Series (1) |
| GSE44614 |
Comparative transcriptomics of Escherichia coli growing in complex environments |
|
