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Sample GSM958143 Query DataSets for GSM958143
Status Public on May 01, 2014
Title BXPC3_NTshRNA_rep1
Sample type RNA
 
Source name BXPC3 NT shRNA, replicate 1
Organism Homo sapiens
Characteristics tissue: Pancreatic cancer cell line
Growth protocol BXPC3 cell lines were plated at 2 million cells/10 cm dish in triplicate and grown RPMI 1640 medium supplemented with 10% heat-inactivated fetal bovine serum. The cultures were incubated for 72 hours at 37 ºC in a humidified incubator with 5% CO2 prior to harvest for RNA isolation.
Extracted molecule total RNA
Extraction protocol Total RNA was isolated from cells with TRIzol Reagent (Life Technologies, Carlsbad, USA) according to manufacturer protocol. RNA was digested with DNase I-Amp Grade (Life Technologies) on-column and purified using an RNeasy Mini kit (QIAGEN, Valencia, USA) according to manufacturer protocols. RNA quality and quantification were performed by analyzing RNA in RNA Nano 6000 chips in a Bioanalyzer2100 following manufacturer protocol (Agilent Technologies, Santa Clara, USA).
Label Cy3
Label protocol Cyanine 3-CTP labeled cRNA target was prepared using One-Color Low Input Quick Amp kit and One Color Spike-In kit (Agilent Technologies) using 200 ng input RNA (RIN>8.0) according to manufacturer protocols. Labeled cRNA was purified using RNeasy Mini kit (QIAGEN) according to manufacturer recommendation. Labeled cRNA was quantified on a NanoDrop ND-1000 UV Spectrophotometer using Microarray/RNA-40 measurement. Specific activity of Cyanine 3 was calculated in pmol /µg with a minimum threshold of 15 pmol/µg.
 
Hybridization protocol 1650 ng of labeled cRNA was fragmented by incubation with 10 µl of 10X Blocking Reagent and 25X Fragmentation Buffer in a 50 µl reaction volume for 30 minutes at 65 °C. 50 µl of 2X GE Hybridization Buffer HI-RPM was added to fragmented cRNA and hybridization mixes then placed onto Agilent Human GE 4X44K v2 microarrays. Hybridization was done for 17 hours at 65 °C by rotating at 10 rpm. Microarray slides were washed in GE Wash Buffer 1 for 2 minutes and pre-warmed GE Wash Buffer 2 for 2 minutes at room temperature.
Scan protocol Slides were scanned immediately after washing on the Agilent DNA Microarray Scanner (G2505B) using one color scan setting for 1x44k array slides (Scan Area 61x21.6 mm, Scan resolution 10um, Dye channel is set to Green and Green PMT is set to 100%).
Description Gene expression
Data processing The scanned images were analyzed with Feature Extraction Software 10 (Agilent) using default parameters to obtain background subtracted and spatially detrended Processed Signal intensities. Features flagged in Feature Extraction as Feature Non-uniform outliers were excluded.
 
Submission date Jul 09, 2012
Last update date May 01, 2014
Contact name Mike Mattie
E-mail(s) mmattie@agensys.com
Organization name Agensys Inc
Department Applied Molecular Biology
Street address 1800 Stewart Street
City Santa Monica
State/province CA
ZIP/Postal code 90404
Country USA
 
Platform ID GPL13497
Series (1)
GSE39207 DDR1 knockdown in BXPC3 cell line

Data table header descriptions
ID_REF
VALUE log2 normalized signal intensity

Data table
ID_REF VALUE
A_23_P146146 -8.57567
A_23_P42935 -1.03795
A_23_P117082 3.6363
A_23_P2683 0.671101
A_24_P358131 3.9062
A_33_P3367647 -8.35762
A_23_P157316 -4.47213
A_32_P14850 3.4586
A_23_P158596 -1.40114
A_23_P350107 0.2212
A_23_P388190 -1.29433
A_23_P106544 3.3626
A_33_P3219745 -8.54358
A_32_P85539 -2.21716
A_23_P94998 -0.6057
A_33_P3235677 -8.54641
A_23_P417014 -8.54797
A_23_P103905 2.1212
A_24_P497186 0.133301
A_23_P118536 -8.62042

Total number of rows: 34127

Table truncated, full table size 722 Kbytes.




Supplementary file Size Download File type/resource
GSM958143_US09473738_252665210504_S01_GE1_107_Sep09_1_2.txt.gz 2.1 Mb (ftp)(http) TXT
Processed data included within Sample table

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