Transcription profiling of human pancreas NS-928 cell lines modulated by COX-2

ABSTRACT: BxPc-3, Miapaca-2 and ASPC-1 cell lines were treated with 100 µM NS-398 in DMSO or with DMSO alone (control) for 48 h. Total-RNA from each sample was isolated with the RNeasy kit of Qiagen (Hilden, Germany) according to the manufacturer's protocol. The integrity of the isolated RNA was checked on an Agilent Bioanalyser 2100 using the RNA 6000 Nano Kit (Agilent Technologies, Palo Alto, USA) according to the manufacturer's protocol as recommended by the manufacturer.
Microarrays were produced and processed as described before. Human cDNAs representing some 7,000 genes that are highly associated with the occurrence of pancreatic cancer – including apoptotic and oncogenic genes, growth factors, angiogenic, cell cycle, metastasis-associated, and housekeeping genes – were PCR-amplified using amino-modified M13 universal primers. PCR-products were purified with Multiscreen PCR (Millipore, Schwalbach, Germany), resuspended in spotting solution (TeleChem International, Sunnyvale, USA) and arrayed onto slides with epoxysilane surface (Quantifoil Micro Tools, Jena, Germany). DNA spotting was done with a Micro-Arrayer of Engineering Services Inc. (Virtek's arrayer system, BioRad, Munich, Germany) using SMP3 pins (TeleChem).

Transcript profiling
Fluorescently labelled cDNA samples were prepared from 15 µg total-RNA and Cy3- or Cy5-labelled dCTP was directly incorporated during first-strand synthesis . Hybridisation to the microarrays was done in hybridisation chambers (TeleChem) at 62°C overnight. After washing in 0.1xSSC, fluorescence signals were detected with a confocal ScanArray 5000 scanner (Packard Bioscience, USA) and analysed with GenePix Pro 6 (Axon Instruments, Union City, USA).

ORGANISM(S): Homo sapiens

SUBMITTER: Mahmoud Youns 

PROVIDER: E-MEXP-1870 | biostudies-arrayexpress |

REPOSITORIES: biostudies-arrayexpress