|
| Status |
Public on Jun 16, 2015 |
| Title |
24h mineralising +/- HDACi replicate 2 |
| Sample type |
RNA |
| |
|
| Channel 1 |
| Source name |
Mineralising control, rat DPC alpha MEM + mineralising stimluli (B-glycerophosphate, Ascorbic acid, Dexamthasone) 37˚C, 5% CO2 24 hrs
|
| Organism |
Rattus norvegicus |
| Characteristics |
strain: Strain Wistar Hann
tissue: dental pulp
agent: no SAHA
|
| Treatment protocol |
Cultures were detached with trypsin/EDTA, homogenized for 30 s using a T10 basic S2-Ultra-Turrax tissue disrupter (IKA, Staufen, Germany)
|
| Growth protocol |
Cells were cultured in supplemenetd alpha MEM in the presence of differentiation stmuli for mineralisation, 37˚C 5% CO2 24h or 14 d.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA was extracted using the RNeasy mini kit (Qiagen, West Sussex, UK). mRNA was isolated on the RNAeasy mini-column assembly (Qiagen)
|
| Label |
Cy3
|
| Label protocol |
mRNA was labelled with Cy5 or Cy3 using the Agilent Two-Color Low RNA input Linear Amplification Kit PLUS
|
| |
|
| Channel 2 |
| Source name |
HDACi, rat DPC alpha MEM + mineralising stimluli (B-glycerophosphate, Ascorbic acid, Dexamthasone) + 1uM SAHA 37˚C, 5% CO2 24 hrs
|
| Organism |
Rattus norvegicus |
| Characteristics |
strain: Strain Wistar Hann
tissue: dental pulp
agent: 1 uM SAHA
|
| Treatment protocol |
Cultures were detached with trypsin/EDTA, homogenized for 30 s using a T10 basic S2-Ultra-Turrax tissue disrupter (IKA, Staufen, Germany)
|
| Growth protocol |
Cells were cultured in supplemenetd alpha MEM in the presence of differentiation stmuli for mineralisation, 37˚C 5% CO2 24h or 14 d.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA was extracted using the RNeasy mini kit (Qiagen, West Sussex, UK). mRNA was isolated on the RNAeasy mini-column assembly (Qiagen)
|
| Label |
Cy5
|
| Label protocol |
mRNA was labelled with Cy5 or Cy3 using the Agilent Two-Color Low RNA input Linear Amplification Kit PLUS
|
| |
|
| |
| Hybridization protocol |
Hybridization and washing of the arrays was carried out using the Agilent Gene Expression Hybridization Kit and Gene Expression Wash Pack according the manufacturer’s instructions.
|
| Scan protocol |
Slides were scanned using the GenePix personal 4100A scanner (Axon) and data was extracted with GenePix Pro 6.1 (Axon).
|
| Description |
Rat DPC alpha MEM + mineralising stimluli (B-glycerophosphate, Ascorbic acid, Dexamthasone) 37˚C, 5% CO2 24 hrs
|
| Data processing |
Spots were flagged absent if the signal was less than background +1 standard deviation in both fluorescent channels. Raw data was exported to GeneSpring GX10 and signals for each replicate spot were background corrected and normalized using Loess normalization. Log2 fluorescence ratios were generated for each replicate spot and averaged. Oligonucleotides were excluded from analysis if >50% of replicates in each condition were flagged absent.
|
| |
|
| Submission date |
Mar 23, 2015 |
| Last update date |
Jun 18, 2015 |
| Contact name |
Hnery Fergus Duncan |
| E-mail(s) |
[email protected]
|
| Phone |
+353 1 612 7356
|
| Organization name |
Dublin Dental University Hospital, Trinity College Dublin
|
| Department |
Restorative Dentistry
|
| Lab |
Cell culture/Materials
|
| Street address |
Lincol Place
|
| City |
Dublin |
| ZIP/Postal code |
Dublin 2 |
| Country |
Ireland |
| |
|
| Platform ID |
GPL14746 |
| Series (1) |
|
