|Public on Apr 01, 2008
|TERT6-217, TERT transfected, Agilent
|human fetal lung fibroblast TIG-1 clone TERT6 transfected with TERT
|human fetal lung fibroblast TIG-1 clone TERT6 at 217 PDL transfected with TERT
|Health Science Research Resources Bank, Tokyo, Japan
|TERT expression vector was transfected to TIG-1 and isolated as a G418 resistant clone. Exponentially growing cultured cells were collected using trypsin followed by rinse with PBS, and freezed until use.
|Cells were cultured in Dulbecco's modified Eagle's minimal essential medium (DMEM; Sigma-Aldrich Japan, Tokyo, Japan) containing 10% heat-inactivated fetal bovine serum (FBS; BioWhittaker, Verviers, Belgium) in a humidified atmosphere of 5% CO2.
|Total RNA was extracted from the frozen cultured cell pellets using QIAGEN RNeasy mini kit (QIAGEN, Inc., Valencia, CA), respectively, according to the manufacturer's protocols. The quality of the RNA was checked using Agilent Technologies 2100 Bioanalyzer (Agilent Technologies, Santa Clara, CA).
|The cDNA was generated from 0.6 microgram of total RNA using reverse transcriptase and a T7 primer using Agilent Low RNA Input Linear Amplification Kit PLUS, One-color (Agilent). cRNA (complementary RNA) was generated via an in vitro transcription reaction using T7 RNA polymerase and cyanine 3-labeled CTP
|The synthesized 1.5 microgram of cRNA, quantified by spectrometry and qualified using an Agilent 2100 Bioanalyzer, was then fragmented and hybridized to each array. After 17hr hybridization, arrays were washed according to the manufacturer's recommended protocol.
|Each microarray was scanned using an Agilent DNA Microarray Scanner.
|Expression levels were quantified by Agilent Feature Extraction software ver. 9.1. and normalized to the median expression value of the whole array spots using GeneSpringTM GX (Agilent Technologies).
|Sep 18, 2007
|Last update date
|Aug 14, 2011
|Research Institute for Radiation Biology and Medicine
|Dept. Translational Cancer Research
|1-2-3 Kasumi, Minami-ku
|Expression profiles of immortal lung fibroblasts