|
| Status |
Public on Aug 23, 2016 |
| Title |
Hcc827-2 |
| Sample type |
RNA |
| |
|
| Source name |
gefitinib-sensitive HCC827cells_rep2
|
| Organism |
Homo sapiens |
| Characteristics |
cell line: Hcc827
tissue: lung adenocarcinoma cells
phenotype: gefitinib-sensitive
|
| Treatment protocol |
To generate a resistant cell line, we exposed HCC827 cells to increasing concentrations of gefitinib Gefitinib concentrations were increased stepwise from 8 to 100 nM when the cells resumed growth kinetics similar to the untreated parental cells. Cells with the ability to grow in 100 nM of gefitinib were obtained 6 months after the initial drug exposure. HCC827-8-1 cell line was isolated as a clone.
|
| Growth protocol |
The cell line was maintained in RPMI-1640 supplemented with 10% FBS (Gibco) at 37°C, with 5% CO2 and humidified atmosphere
|
| Extracted molecule |
total RNA |
| Extraction protocol |
RNA was extracted by Trizol (Life Technology, 15596-018) following the manufacturer's instructions
|
| Label |
cy3
|
| Label protocol |
Cyanine-3 (Cy3) labeled cRNA was prepared from 0.2 ug RNA using the One-Color Low RNA Input Linear Amplification PLUS kit (Agilent) according to the manufacturer's instructions, followed by RNAeasy column purification (QIAGEN, Valencia, CA). Dye incorporation and cRNA yield were checked with the NanoDrop ND-1000 Spectrophotometer.
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| |
|
| Hybridization protocol |
0.6 ug of Cy3-labelled cRNA (specific activity >10.0 pmol Cy3/ug cRNA) was fragmented at 60°C for 30 minutes in a reaction volume of 22.5ul containing 1x Agilent fragmentation buffer and 2x Agilent blocking agent following the manufacturers' instructions. On completion of the fragmentation reaction, 22.5ul of 2x Agilent hybridization buffer was added to the fragmentation mixture and hybridized to OE Human lncRNA Microarray V4.0 (Agilent) for 17 hours at 65°C in a rotating Agilent hybridization oven. After hybridization, microarrays were washed 1 minute at room temperature with GE Wash Buffer 1 (Agilent) and 1 minute with 37°C GE Wash buffer 2 (Agilent), then dried immediately by brief centrifugation.
|
| Scan protocol |
Slides were scanned immediately after washing on the Agilent DNA Microarray Scanner (G2505C) using one color scan setting for 4x180k array slides (Scan Area 61x21.6 mm, Scan resolution 3um, Dye channel is set to Green and Green PMT is set to 100%).
|
| Description |
gene expression in gefitinib-sensitive HCC827 cells
|
| Data processing |
The scanned images were analyzed with Feature Extraction Software 10.7.1.1 (Agilent) using default parameters to obtain background subtracted and spatially detrended Processed Signal intensities as the raw data. Raw data were normalized in quantile algorithm with Genespring 13.0(Agilent). Probe that at least 1 out of 2 samples flagged as Detected were maintained.
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| |
|
| Submission date |
Nov 02, 2015 |
| Last update date |
Aug 23, 2016 |
| Contact name |
Ying Wu |
| E-mail(s) |
wuying-tc@163.com
|
| Phone |
13665186889
|
| Organization name |
Nanjing Medical University Affiliated Cancer Hospital Cancer Institute of Jiangsu Province
|
| Street address |
Baiziting 42
|
| City |
Nanjing |
| ZIP/Postal code |
210009 |
| Country |
China |
| |
|
| Platform ID |
GPL21096 |
| Series (1) |
| GSE74575 |
Microarray expression profile of long noncoding RNAs in EGFR-TKI resistance of lung adenocarcinoma |
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