|
| Status |
Public on May 20, 2010 |
| Title |
1592_1x10^4_vs_1592_1x10^5_rep2 |
| Sample type |
RNA |
| |
|
| Channel 1 |
| Source name |
Right Fore quarter infused with 1x10^4 Staphylococcus aureus mammary tissue
|
| Organism |
Bos taurus |
| Characteristics |
Dose: 1x10^4
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Approximately 3 g of frozen mammary tissue was placed in liquid nitrogen and pulverised with a hammer followed by the addition of 25 ml of Trizol reagent (Invitrogen). The tissue was homogenised using an Ultraturrex probe (IKA, 3 cycles of 2 seconds). RNA was then extracted from three equivalent samples from each udder quarter according to the Trizol manufacturer.s protocol (Invitrogen) and 1 mg aliquots were stored as pellets under ethanol at -80.C. RNA was further purified using an RNeasy Midi Kit (Qiagen) and each sample was treated twice with DNase I on column (Qiagen DNase I) and after elution (Ambion DNase I) to eliminate any contaminating genomic DNA.
|
| Label |
Cy5
|
| Label protocol |
cDNA prepared from each sample using 20 ug of total RNA per dye channel per microarray, was reverse transcribed with Superscript III (Invitrogen) in the presence of 2-aminoallyl-dUTP (Sigma Chemical Co.) using both oligo-dT18 (2 .g) and pd(N)6 random hexamer (1 .g) (Amersham Bioscience, UK) to prime cDNA synthesis. First strand cDNAs were purified and subsequently labelled using n-hydroxysuccinate (NHS)-derivatized Cy3 and Cy5 dyes (Amersham Biosciences, UK)
|
| |
|
| Channel 2 |
| Source name |
Left Hind quarter infused with 1x10^5 Staphylococcus aureus mammary tissue
|
| Organism |
Bos taurus |
| Characteristics |
Dose: 1x10^5
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Approximately 3 g of frozen mammary tissue was placed in liquid nitrogen and pulverised with a hammer followed by the addition of 25 ml of Trizol reagent (Invitrogen). The tissue was homogenised using an Ultraturrex probe (IKA, 3 cycles of 2 seconds). RNA was then extracted from three equivalent samples from each udder quarter according to the Trizol manufacturer.s protocol (Invitrogen) and 1 mg aliquots were stored as pellets under ethanol at -80.C. RNA was further purified using an RNeasy Midi Kit (Qiagen) and each sample was treated twice with DNase I on column (Qiagen DNase I) and after elution (Ambion DNase I) to eliminate any contaminating genomic DNA.
|
| Label |
Cy3
|
| Label protocol |
cDNA prepared from each sample using 20 ug of total RNA per dye channel per microarray, was reverse transcribed with Superscript III (Invitrogen) in the presence of 2-aminoallyl-dUTP (Sigma Chemical Co.) using both oligo-dT18 (2 .g) and pd(N)6 random hexamer (1 .g) (Amersham Bioscience, UK) to prime cDNA synthesis. First strand cDNAs were purified and subsequently labelled using n-hydroxysuccinate (NHS)-derivatized Cy3 and Cy5 dyes (Amersham Biosciences, UK)
|
| |
|
| |
| Hybridization protocol |
Hybridisation was performed at 44°C in a 50% formamide hybridisation buffer for 16 h. Following hybridisation, three washes were applied to the arrays: 2 x SSC, 0.1% SDS for 15 min, pre-warmed to 44°C; 0.2 x SSC for 15 min; and 0.06 x SSC for 5 min. Slides were washed once in high purity water to remove any remaining salt and dried in a centrifuge at 40xg for 5 min.
|
| Scan protocol |
Dried slides were scanned immediately using the GenePix. 4000 array scanner. GenePix.Pro software version 5.0 (Axon Instruments Inc.) was then used to process array images, align spots, integrate robot-spotting files with the microarray image, and export reports of spot intensity data. Slides were visually examined and spots with irregular morphology were excluded from data analysis
|
| Description |
Bovine mammary tissue
|
| Data processing |
After background correction and removal of flagged values, log base 2 expression ratios were mean centered and linear transformed to obtain the log and linear values given in the data table
|
| |
|
| Submission date |
Nov 26, 2007 |
| Last update date |
Aug 14, 2011 |
| Contact name |
Ylva Strandberg-Lutzow |
| E-mail(s) |
[email protected]
|
| Organization name |
CSIRO
|
| Department |
Livestock Industries
|
| Street address |
306 Carmody Road
|
| City |
St Lucia |
| State/province |
Queensland |
| ZIP/Postal code |
4067 |
| Country |
Australia |
| |
|
| Platform ID |
GPL6082 |
| Series (2) |
| GSE9680 |
Response of bovine mammary tissue to experimental Staphylococcus aureus infection experiment 2. |
| GSE9685 |
Response of bovine mammary tissue to experimental Staphylococcus aureus infection |
|
| Data table header descriptions |
| ID_REF |
|
| VALUE |
Base-2 logarithm of channel 1 corrected median signal divided by channel 2 corrected median signal with flagged values removed |
| F635 Median |
Channel 1 median intensity |
| F635 Mean |
Channel 1 mean intensity |
| F635 SD |
Channel 1 standard deviation |
| B635 Median |
Channel 1 median background intensity |
| B635 Mean |
Channel 1 mean background intensity |
| B635 SD |
Channel 1 background standard deviation |
| F532 Median |
Channel 2 median intensity |
| F532 Mean |
Channel 2 mean intensity |
| F532 SD |
Channel 2 standard deviation |
| B532 Median |
Channel 2 median background intensity |
| B532 Mean |
Channel 2 mean background intensity |
| B532 SD |
Channel 2 background standard deviation |
| Ratio of Medians |
Unnormalized, untransformed ratio of medians |
| Ratio of Means |
Unnormalized, untransformed ratio of means |
| Median of Ratios |
Unnormalized, untransformed median of ratios |
| Mean of Ratios |
Unnormalized, untransformed mean of ratios |
| Flags |
0 denotes satisfactory features, while <0 denotes features that did not meet criteria |
| UNF_VALUE |
Base-2 logarithm of channel 1 corrected median signal divided by channel 2 corrected median signal |
