|
| Status |
Public on Aug 13, 2020 |
| Title |
Meniscus_24W_OA7 |
| Sample type |
RNA |
| |
|
| Source name |
Meniscus, 24W, OA7
|
| Organism |
Sus scrofa |
| Characteristics |
tissue: meniscus
gender: male
age: 6 to 7 months
group: experimental
|
| Growth protocol |
Cryopulverization of tissue using the Biopulverizer
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Take an appropriate amount (50-100mg) of properly preserved tissue samples, use BioPulverizerTM to freeze and crush the tissue, add 1ml of RNA extraction reagent TRIzol (Invitrogen), use Mini-Bead-Beater-16 to homogenize and extract RNA. Use NanoDrop-1000 spectrophotometer to quantify RNA, and perform denaturing agarose gel electrophoresis with formaldehyde electrophoresis reagent to detect RNA purity and integrity.
|
| Label |
Cy3
|
| Label protocol |
Total RNA from each sample was linearly amplified and labeled with Cy3-UTP. The Labeled cRNAs were purified by RNeasy Mini Kit (Qiagen). The concentration and specific activity of the labeled cRNAs (pmol Cy3/μg cRNA) were measured by NanoDrop ND-1000.
|
| |
|
| Hybridization protocol |
1 μg of each labeled cRNA was fragmented by adding 11 μl 10 × Blocking Agent and 2.2 μl of 25×Fragmentation Buffer, then heated at 60°C for 30 min, and finally 55 μl 2 × GE Hybridization buffer was added to dilute the labeled cRNA. 100 μl of hybridization solution was dispensed into the gasket slide and assembled to the gene expression microarray slide. The slides were incubated for 17 hours at 65°C in an Agilent Hybridization Oven. The hybridized arrays were washed, fixed and scanned with using the Agilent DNA Microarray Scanner (part number G2505C).
|
| Scan protocol |
Agilent Feature Extraction software (version 11.0.1.1) was used to analyze acquired array images.
|
| Description |
After 24 weeks after the pig underwent anterior cruciate ligament surgery, the meniscus tissue was taken to extract RNA, and then gene chip detection was performed
|
| Data processing |
Quantile normalization and subsequent data processing were performed with using the GeneSpring GX v12.1 software package (Agilent Technologies). After quantile normalization of the raw data, genes that at least 7 out of 14 samples have flags in Detected (“All Targets Value”) were chosen for further data analysis. Differentially expressed genes with statistical significance between the two groups were identified through Volcano Plot filtering. Differentially expressed genes between the two samples were identified through Fold Change filtering. Hierarchical Clustering was performed using the R scripts. GO analysis and Pathway analysis were performed in the standard enrichment computation method.
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| |
|
| Submission date |
Aug 12, 2020 |
| Last update date |
Aug 13, 2020 |
| Contact name |
Hui Huang |
| Organization name |
Hainan Provincial People's Hospital
|
| Street address |
19 Xiuhua Road, Xiuying District
|
| City |
Haikou |
| ZIP/Postal code |
570311 |
| Country |
China |
| |
|
| Platform ID |
GPL16571 |
| Series (1) |
| GSE156132 |
Bioinformatics Analysis of Degenerative Meniscus Tissue in Knee Osteoarthritis of Wuzhishan Pig |
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