|
| Status |
Public on Jan 12, 2013 |
| Title |
PLC_PTvsLM |
| Sample type |
RNA |
| |
|
| Channel 1 |
| Source name |
PLC-PT
|
| Organism |
Homo sapiens |
| Characteristics |
cell type: Primary human HCC tumor cell line
cell line (derived): PLC8024
|
| Growth protocol |
All cell lines are maintain at 37oC incubator with 5% CO2 with DMEM supplement with 10% FBS
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA were extracted by Trizol reagent accroding to manufacturer's protocol
|
| Label |
Cy3
|
| Label protocol |
Each total RNA sample, 1μl of NCode™ Multi-Species miRNA Microarray Controls (2fmol/ul) was spiked, and Poly (A) tailing reactions were set up following the protocol recommended in the NCode™ Rapid miRNA Labeling System Manual. The Poly (A) tailed reactions were ligated to labeled DNA polymers by using the 6X Alexa Fluor® 3 (A3) Rapid Ligation Mix or 6X Alexa Fluor® 5 (A5) Rapid Ligation Mix following the protocol recommended in the NCode™ Rapid miRNA Labeling System Manual.
|
| |
|
| Channel 2 |
| Source name |
PLC-LM
|
| Organism |
Homo sapiens |
| Characteristics |
cell type: Metastatic human HCC tumor cell line
cell line (derived): PLC8024
|
| Growth protocol |
All cell lines are maintain at 37oC incubator with 5% CO2 with DMEM supplement with 10% FBS
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA were extracted by Trizol reagent accroding to manufacturer's protocol
|
| Label |
Cy5
|
| Label protocol |
Each total RNA sample, 1μl of NCode™ Multi-Species miRNA Microarray Controls (2fmol/ul) was spiked, and Poly (A) tailing reactions were set up following the protocol recommended in the NCode™ Rapid miRNA Labeling System Manual. The Poly (A) tailed reactions were ligated to labeled DNA polymers by using the 6X Alexa Fluor® 3 (A3) Rapid Ligation Mix or 6X Alexa Fluor® 5 (A5) Rapid Ligation Mix following the protocol recommended in the NCode™ Rapid miRNA Labeling System Manual.
|
| |
|
| |
| Hybridization protocol |
The two differentially labeled reactions were combined into one tube and the volume was reduced by half in a SpeedVac® Concentrator. BSA (50mg/ml) was then added to a total volume of 28.5ul. The samples were incubated with 28.5ul of 2X Enhanced hybridization buffer at 65oC for 10 min and then loaded onto NCode™ Multi-Species miRNA Microarrays V2. The arrays were mounted with the Maui Mixer SL and hybridized overnight (16-20 h) at 52oC with constant mixing.
|
| Scan protocol |
The washes were performed following the standard protocol recommended in the NCode™ Rapid miRNA Labeling System Manual, and the arrays were scanned using a GenePix® 4000B microarray scanner (Molecular Devices).
|
| Description |
Analysis carried out by comparing the primary tumor cell line established from primary liver tumor in mice and its corresponding lung metastatic tumor cell line
|
| Data processing |
The scanned array images were annotated and analyzed using the GenePix software and the .GAL files containg the array list name for the human samples.Analysis was performed by Microsoft Excel
|
| |
|
| Submission date |
Jan 11, 2013 |
| Last update date |
Jan 12, 2013 |
| Contact name |
Wing Lung Simon Yau |
| E-mail(s) |
swlyau@hku.hk
|
| Phone |
(852)28199622
|
| Fax |
(852)28199634
|
| Organization name |
The University of Hong Kong
|
| Department |
Surgery
|
| Lab |
L9-29
|
| Street address |
21 Sassoon Road
|
| City |
Pokfulam, Hong Kong |
| ZIP/Postal code |
NA |
| Country |
Hong Kong |
| |
|
| Platform ID |
GPL16496 |
| Series (1) |
| GSE43445 |
MicroRNA profiling in primary and metastatic HCC cell lines using PLC8024 and MHCC97H derived cell lines |
|
