|
| Status |
Public on Jul 03, 2014 |
| Title |
MM231-D3H2LN-GFP cells |
| Sample type |
RNA |
| |
|
| Source name |
breast cancer cell line
|
| Organism |
Homo sapiens |
| Characteristics |
cell type: MM231-D3H2LN-GFP cells
|
| Treatment protocol |
We sorted MM231-D3H2LN-GFP-BM2 cells co-cultured with BM-MSCs into CD44- and CD44+ populations.
|
| Growth protocol |
To track metastasis and establish a bone marrow-metastatic cell line, lentiviral infection of MDA-MB-231-luc-D3H2LN cells, was performed with the pGreenPuro Scramble Hairpin Control (construct MZIP000-PA-1, System Biosciences) to establish breast cancer cells that stably expressed both firefly luciferase and a GFP cloned from copepod Pontellina plumata (cop-GFP, also known as ppluGFP2). These cells, which we called MDA-MB-231-luc-D3H2LN-GFP, were cultured in RPMI containing 10% heat-inactivated fetal bovine serum (FBS) and penicillin with 100 units/mL, streptomycin with 100 µg/mL, and amphotericin B with 0.25µg/mL at 37°C in 5% CO2. The left heart ventricle of eight-week-old female C.B-17/Icr-scid/scidJc1 mice were injected with 1 × 105 MDA-MB-231-luc-D3H2LN-GFP cells suspended in 100 μl of PBS. The subsequent development of metastasis was monitored by injecting luciferin into the mice and measuring bioluminescence using an IVIS imaging system and Living Image 2.50 analysis software. On day 10, bioluminescence was detected in the bilateral legs of a mouse that had received an intracardiac injection. The mouse was sacrificed, and bone marrow was extracted from the legs, cultured in RPMI as described for the parental MDA-MB-231-luc-D3H2LN-GFP cell line above, and microscopically analyzed for GFP-positive cells. An MDA-MB-231-luc-D3H2LN-GFP cell was isolated and cultured under zeocin selection to establish clones, which we called MM231-D3H2LN-GFP-BM2 cells, cultured in RPMI as described above for the parental line.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA was extracted from cultured cells using the QIAzol reagent and the miRNeasy Mini Kit (Qiagen) following the manufacturer's recommendations.
|
| Label |
Cy3
|
| Label protocol |
Agilent one-color Low RNA Input Linear Amplification Kit labeling protocol
|
| |
|
| Hybridization protocol |
Agilent one-color gene expression hyb/wash protocol
|
| Scan protocol |
Microarray slides were scanned in an Agilent Technologies G2505C Microarray Scanner at 3 micron resolution.
|
| Description |
Gene expression of the parental cells
|
| Data processing |
The scanned images were analyzed with Feature Extraction Software 10.7.3.1 (Agilent) using default parameters (protocol GE1_107_Sep09 and Grid: 039494_D_F_20120628) to obtain background subtracted and spatially detrended Processed Signal intensities. Features flagged in Feature Extraction as Feature Non-uniform outliers were excluded.
|
| |
|
| Submission date |
May 23, 2014 |
| Last update date |
Apr 23, 2018 |
| Contact name |
Yusuke Yoshioka |
| E-mail(s) |
yyoshiok@ncc.go.jp
|
| Organization name |
National Cancer Center Research Institute
|
| Department |
Division of Molecular and Cellular Medicine
|
| Street address |
5-1-1, Tsukiji, Chuo-ku
|
| City |
Tokyo |
| ZIP/Postal code |
104-0045 |
| Country |
Japan |
| |
|
| Platform ID |
GPL16699 |
| Series (1) |
| GSE57921 |
Gene expression signatures for breast cancer cells metastatic to bone marrow |
|
| Relations |
| Reanalyzed by |
GSE113533 |
