|
| Status |
Public on Dec 31, 2010 |
| Title |
SC_T_60min_rep2 |
| Sample type |
RNA |
| |
|
| Channel 1 |
| Source name |
Yeast RNA, HMF treated, 60min
|
| Organism |
Saccharomyces cerevisiae |
| Characteristics |
strain: NRRL Y-12632
|
| Treatment protocol |
HMF was added into the culture at a final concentration of 30 mM 6 h after the inoculation. Cultures grown under the same conditions without the HMF treatment served as a control.
|
| Growth protocol |
The yeast was maintained and cultured on a synthetic complete (SC) medium.Culture inocula were prepared using freshly grown cells harvested at logarithmic growth phase after incubation with agitation of 250 rpm at 30˚C for 16 h. Cells were incubated on SC medium in a fleaker fermentation system at 30˚C with agitation.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA was extracted by hot phenol method and further purified using RNeasy Mini Kit following manufacturer's instructions
|
| Label |
Cy3
|
| Label protocol |
Fifteen μg of total RNA was used for each sample labeling reaction in a total volume of 30 μl. RNA probe together with incorporated RNA controls was labeled using an indirect dUTP Cy3 or Cy5 dye as described by Liu and Slininger (2007).
|
| |
|
| Channel 2 |
| Source name |
Yeast RNA, HMF untreated, 0min
|
| Organism |
Saccharomyces cerevisiae |
| Characteristics |
strain: NRRL Y-12632
|
| Treatment protocol |
HMF was added into the culture at a final concentration of 30 mM 6 h after the inoculation. Cultures grown under the same conditions without the HMF treatment served as a control.
|
| Growth protocol |
The yeast was maintained and cultured on a synthetic complete (SC) medium.Culture inocula were prepared using freshly grown cells harvested at logarithmic growth phase after incubation with agitation of 250 rpm at 30˚C for 16 h. Cells were incubated on SC medium in a fleaker fermentation system at 30˚C with agitation.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA was extracted by hot phenol method and further purified using RNeasy Mini Kit following manufacturer's instructions
|
| Label |
Cy5
|
| Label protocol |
Fifteen μg of total RNA was used for each sample labeling reaction in a total volume of 30 μl. RNA probe together with incorporated RNA controls was labeled using an indirect dUTP Cy3 or Cy5 dye as described by Liu and Slininger (2007).
|
| |
|
| |
| Hybridization protocol |
Hybridization was performed based on Hegde et al (2000) with modifications using HS 4800 Hybridization station.
|
| Scan protocol |
scanned on a genePix 4000b scanner. A pre-scan control mini-array was used to adjust PMT Gain against Cy3 and Cy5 channels and the ratios of signal intensities between Cy3 and Cy5 were balanced to 1.0 using the calibration control as described by Liu and Slininger (2007). Each spot was individually examined and adjusted or flagged if necessary.
|
| Description |
Biological replicate 2 of 2. HMF (30mM) treated, harvested at 60min.
|
| Data processing |
All test samples (ch1) were compared to the same reference. Raw data file for the reference are available as a supplementary file on the Series record.
Median of foreground signal intensity subtracted by background for each dye channel was used for analysis. Raw data for each slide were normalized based on spike-in control gene CAB. GeneSpring GX 10.0 (Agilent software) was used. expression values less than 100 in 7 of 16 samples were filtered out from probesets, then a 2-way ANOVA analysis (p≤ 0.05) was performed. Genes showing statistically significant differential expressions with a minimum of 2-fold changes were selected for Principal Component Analysis and clustering analysis by Hierarchical and Self Organizing Maps.
|
| |
|
| Submission date |
Jul 14, 2010 |
| Last update date |
Dec 31, 2010 |
| Contact name |
Z Lewis Liu |
| E-mail(s) |
[email protected]
|
| Organization name |
USDA-ARS
|
| Lab |
NCAUR
|
| Street address |
1815 N University
|
| City |
Peoria |
| State/province |
IL |
| ZIP/Postal code |
61604 |
| Country |
USA |
| |
|
| Platform ID |
GPL10684 |
| Series (1) |
| GSE22939 |
Comparative transcriptome profiling analyses during the lag phase uncover YAP1, PDR1, PDR3, RPN4 and HSF1 as key regulatory genes in genomic adaptation to the lignocellulose derived inhitibor-stress for saccharomyces cerevisiae |
|
