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Sample GSM1532618 Query DataSets for GSM1532618
Status Public on Oct 28, 2014
Title CAF32_control_rep2
Sample type RNA
 
Source name CAF32_control_, replicate 2
Organism Homo sapiens
Characteristics tissue: stomach
cell type: gastric cancer-associated fibroblast (CAF)
Treatment protocol Fresh gastric cancer tissues were obtained from 2 patients and minced with scalpels in a culture dish. Samples were enzymatically dissociated in 20 ml of D/F12+serum media containing collagenase I in a 37℃ incubation for 12-15 h. After digestion, samples were centrifuged at 700 rpm for 5 min to separate epithelial cells and fibroblast cells. Cancer-associated fibroblast cells (named as CAF14 and CAF32 from two gastric cancer tissues, respevtively) were collected from the supernatant by centrifugation at 800 rpm for 8 min, washed twice with PBS, and cultured in D/F12 media supplemented with 10% FBS and 1% antibiotics.
Extracted molecule total RNA
Extraction protocol RNA was extracted using RNeasy kit from 2 cancer-associated fibroblasts obtained by treatment protocol. RNA was quantified using a NanoDrop-1000 spectrophotometer and quality was monitored with the Agilent 2100 Bioanalyzer (Agilent Technologies, Santa Clara, CA).
Label Cy3
Label protocol 0.5 ug of total RNA was amplified and labeled using Agilent Quick Amp Labeling Kit. After labeling, cRNA was purified with Qiagen’s RNeasy Mini-spin columns (QIAGEN, Germantown, MD) and quantified using NanoDrop ND-1000 UV-VIS Spectrophotometer for cyanine 3 dye concentration, RNA absorbance ratio (260 nm/280 nm), and cRNA concentration.
 
Hybridization protocol Cyanine 3-labled linearly amplified cRNA was mixed with 10X blocking agent, nuclease-free water, and 25X fragmentation buffer provided in the Agilent Gene Expression Hybridization Kit (Agilent Technologies), and incubated for 30 min at 60oC to fragment RNA. After adding 2X GEx hybridization buffer Hi-RPM, the amplified fragmented RNA was hybridized on a Agilent SurePrint G3 Human GE v2 8x60K Microarray Chip using a SureHyb DNA Microarray Hybridization Chamber in a DNA Microarray Hybridization Oven (Agilent Technologies) at 10 rpm, 65℃ for 17 hours.
Scan protocol Slides were scanned with an Agilent DNA microarray scanner (Agilent Microarray Scanner–G2565BA, Agilent Technologies).
Description Gene expression
Data processing Feature Extraction Software v9 (Agilent Technologies) was used to extract and analyze the signals. To validate the amplification process, amplified and unamplified RNAs were labeled, hybridized and compared.
 
Submission date Oct 27, 2014
Last update date Oct 28, 2014
Contact name Chang Ohk Sung
Organization name Asan Medical Center
Department Pathology
Street address Song-Pa gu,Pung-nap dong
City Seoul
ZIP/Postal code 05505
Country South Korea
 
Platform ID GPL13607
Series (2)
GSE62738 Twist1 knockdown in two gastric cancer-associated fibroblast lines
GSE62740 Twist1 is a key regulator for cancer-associated fibroblasts

Data table header descriptions
ID_REF
VALUE Normalized signal intensity after quantile normalization method using BRBarry tool.

Data table
ID_REF VALUE
1 17.60308456
2 5.685267448
3
4 9.777822495
5 11.42987156
6 7.032653332
7 13.01953888
8 11.4824791
9 5.573019505
10 7.357155323
11 5.294849873
12 10.8462019
13 11.20734787
14 10.53378296
15 13.60234451
16 5.445006371
17 9.622776985
18 7.35205698
19 5.978158474
20 10.71633816

Total number of rows: 62976

Table truncated, full table size 872 Kbytes.




Supplementary file Size Download File type/resource
GSM1532618_CA32_ShNs_2nd.txt.gz 3.1 Mb (ftp)(http) TXT
Processed data included within Sample table

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