|
| Status |
Public on May 17, 2013 |
| Title |
side population-PDLCs |
| Sample type |
RNA |
| |
|
| Source name |
side population of human periodontal ligament cells
|
| Organism |
Homo sapiens |
| Characteristics |
cell type: side population of human periodontal ligament cells
|
| Extracted molecule |
total RNA |
| Extraction protocol |
total RNA was collected with RNAzol RT (Molecular Research Center, OH, USA). A total of 250 ng of total RNA from MP cells was used for microRNA array. In the case of SP cells, since there was not satisfactory amount of RNA due to the extremely low number of sorted cells (0.1%), a total of 3 μL was applied for the array.
|
| Label |
Hy3
|
| Label protocol |
（miRCURY LNA microRNA Array Hi-Power Labeling kit） (Mixture: 1 rxn.) ------------------------------- Labeling Buffer: 3 μL DMSO: 2 μL Labeling enzyme: 1 μL ------------------------------- Total: 6 μL Add CIP reaction: 5 μL Hy3 Fluorescent label: 1.5 μL Incubate at 16 ℃ for 2 hr, at 65 ℃ 15 min.
|
| |
|
| Hybridization protocol |
Denature the solution at 95 ℃ for 5 min. Centrifuge at 12,000 g at 4 ℃ for 2 min. Add 30 μL of 20 x salt Buffer to ditch of “Hybridization Chamber”. Slide is capped by “MASTUNAMI Gap Cover Glass (CG00044 or 24*60 25PCS)”. Fill the gap with 45 μL of the solution. Incubate in 56 ℃ water bath for 16-20 hr. <Day2> Wash Separate gap cover glass from the slide in “Wash buffer A” (pre-heated at 56 ℃). Wash with “Wash buffer A” (pre-heated at 56 ℃) for 2 min. Wash briefly (one plunge) with “Wash buffer B”. Wash with “Wash buffer B” for 2 min. Wash with “Wash buffer C” for 2 min. Centrifuge at 1000 rpm for 5 min.
|
| Scan protocol |
Scanner; G2505C (Agilent) Software; Agilent Scan Control (Agilent) Green and Red PMT: 100 %, Scan resolution: 10 um,
|
| Description |
enriched in stell cells
|
| Data processing |
Software; Feature Extraction 10.7.3.1 (Agilent) ・Probe Name, ・Gene Name, ・gProcessed Signal (Hy3), Calculate the average of replicated spots (Hy3 data). Average of signal intensity in 4 repeated spots in the array was considered as the final data for each probe.
|
| |
|
| Submission date |
May 16, 2013 |
| Last update date |
May 17, 2013 |
| Contact name |
Emilio Satoshi Hara |
| E-mail(s) |
gmd421209@s.okayama-u.ac.jp
|
| Organization name |
Okayama University
|
| Street address |
Shikata-cho 2-5-1
|
| City |
Okayama-shi |
| State/province |
Okayama-ken |
| ZIP/Postal code |
700-8558 |
| Country |
Japan |
| |
|
| Platform ID |
GPL11434 |
| Series (1) |
| GSE47025 |
miRNA-720 controls stem cell phenotype, proliferation and differentiation of dental pulp cells |
|
