|
| Status |
Public on Apr 16, 2014 |
| Title |
A4 Replicate 2 |
| Sample type |
RNA |
| |
|
| Channel 1 |
| Source name |
A4, osmotolerant mutant
|
| Organism |
Escherichia coli |
| Characteristics |
genotype/variation: Adaptive mutant
population: A4
average fitness: 0.434286415079969
|
| Growth protocol |
Grown in glucose M9 media supplemented with 0.55 M NaCl
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Samples were grown to mid exponential phase (OD approx 0.5). Cultures were then harvested with rapid filtration and resuspended in RNALater to stop RNA degradation. The Zymo Quick-RNA miniprep kit was used to extract pure RNA for subsequent analysis
|
| Label |
Cy3
|
| Label protocol |
RNA was converted to amino-allyl cDNA using Superscript III RT, purified by ethanol precipitation, and resuspended in dye coupling buffer. Cy3 and Cy5 were utilized to label the experimental samples and reference as specified by GE Amersham.
|
| |
|
| Channel 2 |
| Source name |
2xOriT
|
| Organism |
Escherichia coli BW25113 |
| Characteristics |
strain: unevolved parent strain
strain description: E. coli BW25113 with F oriTs integrated at mbhA and hyfC
|
| Growth protocol |
Grown in glucose M9 media supplemented with 0.55 M NaCl
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Samples were grown to mid exponential phase (OD approx 0.5). Cultures were then harvested with rapid filtration and resuspended in RNALater to stop RNA degradation. The Zymo Quick-RNA miniprep kit was used to extract pure RNA for subsequent analysis
|
| Label |
Cy5
|
| Label protocol |
RNA was converted to amino-allyl cDNA using Superscript III RT, purified by ethanol precipitation, and resuspended in dye coupling buffer. Cy3 and Cy5 were utilized to label the experimental samples and reference as specified by GE Amersham.
|
| |
|
| |
| Hybridization protocol |
Microarray samples were hybridized and washed according to the manufacturer's instructions for two color prokaryotic microarrays.
|
| Scan protocol |
Slides were scanned with an Axon 4200A GenePix Scanner at 5um resolution. PMT was equal to 600 for both dye channels
|
| Description |
Fitness Calculation: Mutant growth rate / WT growth rate - 1
Conditions: 0.65 M NaCl, minimal media + glucose (0.5% w/v) + 50 ug/ml tryptophan
Measurement: 96 well plate, TECAN Infinite M200
|
| Data processing |
Data were normalized using the LOWESS algorithm and analyzed for statistical significance using the MeV package
|
| |
|
| Submission date |
Oct 23, 2013 |
| Last update date |
Apr 16, 2014 |
| Contact name |
James Winkler |
| E-mail(s) |
jdwinkler@tamu.edu
|
| Organization name |
Texas A&M University
|
| Lab |
Kao Lab
|
| Street address |
3122 TAMU
|
| City |
College Station |
| State/province |
TX |
| ZIP/Postal code |
77843 |
| Country |
USA |
| |
|
| Platform ID |
GPL13359 |
| Series (1) |
| GSE51611 |
Unravelling osmotololerance in Escherichia coli using a pseudosexual model system |
|
