|
| Status |
Public on Dec 17, 2007 |
| Title |
6_040227 Hog3+Hog4 Hog3cy5+Hog4cy3 p3cy5 2419 test-ck |
| Sample type |
RNA |
| |
|
| Channel 1 |
| Source name |
control
|
| Organism |
Sus scrofa |
| Characteristics |
The RNA was isolated from back fat tissue of Chinese Miniature pigs, which was 3 month-old hog.
|
| Treatment protocol |
The pig was fed without clenbuterol.
|
| Growth protocol |
Eight Chinese miniature pigs were used in the experiments. Four hogs and four sows, all 4 weeks old, were housed in the Nutrition and Metabolism Laboratory at the China Agriculture University. They were raised under exactly the same conditions and were fed the same diets until 8 weeks (average body weight 17 kg). They were randomly divided into 4 groups with each group having two pigs with the same gender and the same parents. For the following 4 weeks, one pig in each group was fed 25 mg/kg clenbuterol twice daily in their diets as the test pig, while the other was fed the same diet without clenbuterol as the control. Then one group of hogs and one group of sows were slaughtered for analysis. These two groups are referred to as the 3 month-old pigs. The other two groups were fed with/without 50 mg/kg clenbuterol twice daily in their diets for another 4 weeks and slaughtered for analysis. These two groups are referred to as the 4 month-old pigs. Approximately 1 g biopsies were taken from the back fat adipose tissues (at the fifth lumbar vertebra level) of each pig. The adipose tissue samples were washed in sterile water, snap frozen in liquid nitrogen and stored at -80℃.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA of the adipose tissue was extracted with TRIZOL reagent (Invitrogen, Gaithersburg, MD, USA) and further purified with an RNeasy mini kit (Qiagen, Valencia, CA, USA) according to the manufacturers instructions.
|
| Label |
cy3
|
| Label protocol |
Reverse transcription was done with 10 µg total RNA as the template to synthesize cDNA incorporated with flourence dye Cy3-dCTP or Cy5-dCTP. Probes were purified on a Qiagen spin column (Qiagen, Valencia, CA, USA). Mix dye-labeled cDNA together and dry dye-labeled cDNA were used for hybridization.
|
| |
|
| Channel 2 |
| Source name |
test
|
| Organism |
Sus scrofa |
| Characteristics |
The RNA was isolated from back fat tissue of Chinese Miniature pigs, which was 3 month-old hog.
|
| Treatment protocol |
The pig was fed with clenbuterol.
|
| Growth protocol |
Eight Chinese miniature pigs were used in the experiments. Four hogs and four sows, all 4 weeks old, were housed in the Nutrition and Metabolism Laboratory at the China Agriculture University. They were raised under exactly the same conditions and were fed the same diets until 8 weeks (average body weight 17 kg). They were randomly divided into 4 groups with each group having two pigs with the same gender and the same parents. For the following 4 weeks, one pig in each group was fed 25 mg/kg clenbuterol twice daily in their diets as the test pig, while the other was fed the same diet without clenbuterol as the control. Then one group of hogs and one group of sows were slaughtered for analysis. These two groups are referred to as the 3 month-old pigs. The other two groups were fed with/without 50 mg/kg clenbuterol twice daily in their diets for another 4 weeks and slaughtered for analysis. These two groups are referred to as the 4 month-old pigs. Approximately 1 g biopsies were taken from the back fat adipose tissues (at the fifth lumbar vertebra level) of each pig. The adipose tissue samples were washed in sterile water, snap frozen in liquid nitrogen and stored at -80℃.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA of the adipose tissue was extracted with TRIZOL reagent (Invitrogen, Gaithersburg, MD, USA) and further purified with an RNeasy mini kit (Qiagen, Valencia, CA, USA) according to the manufacturers instructions.
|
| Label |
cy5
|
| Label protocol |
Reverse transcription was done with 10 µg total RNA as the template to synthesize cDNA incorporated with flourence dye Cy3-dCTP or Cy5-dCTP. Probes were purified on a Qiagen spin column (Qiagen, Valencia, CA, USA). Mix dye-labeled cDNA together and dry dye-labeled cDNA were used for hybridization.
|
| |
|
| |
| Hybridization protocol |
See Pronto Universal hybridization kit Quick Reference Guide.
|
| Scan protocol |
Arrays were scanned with a ScanArray Express scanner (Parckard Bioscience, Kanata, OT，USA) with the obtained images analyzed with GenePix Pro 4.0 (Axon Instruments, Foster City, CA ) and Acuity 4.0 (Axon Instruments, Foster City, CA).
|
| Description |
To avoid dye bias, the experiments were performed in duplicate by dye swap with Cy5-dCTP first used with the test pig and Cy3-dCTP used with the control pig and then Cy3-dCTP used with the test pig and Cy5-dCTP used with the control pig.
|
| Data processing |
The resulting microarray data was then normalized using the space and intensity-dependent normalization in the LOWESS program. Each gene was represented in triplicate on each slide. The intensity (median) of each spot was analyzed using the student t-test to identify the differentially expressed genes (P<0.05). Low quality spots were filtered out before the student t-test analysis. Low spots refer to stained spots with bad images or spots with the intensities (median) lower than 200 (too weak) or greater than 60000 (saturated).
|
| |
|
| Submission date |
Jun 10, 2007 |
| Last update date |
Aug 14, 2011 |
| Contact name |
Jin Zhang |
| E-mail(s) |
zhangjin7688@163.com
|
| Organization name |
China Agricultural University
|
| Street address |
2#, Yuanmingyuan West Road
|
| City |
Beijing |
| ZIP/Postal code |
100094 |
| Country |
China |
| |
|
| Platform ID |
GPL5374 |
| Series (1) |
| GSE8093 |
Differential Gene Expression Profile in Pig Adipose Tissue Treated with/without Clenbuterol |
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