Notch targets in DmD8 and KC Drosophila cells

To identify genes upregulated in response to Notch signalling in DmD8 cells. To identify genes upregulated in response to Notch signalling in KC cells. Keywords: Expression analysis at a single timepoint (30' after Notch activation) DmD8 cells were obtained from the Drosophila Genomics Resource Center (http://dgrc.cgb.indiana.edu). In the first experiment, 'control cells versus Notch activated cells' mRNA was extracted from DmD8 cells incubated in the absence or presence of EDTA to activate Notch (cells were harvested 30 minutes after addition of EDTA). In a second experiment, 'Notch activated in presenilin inhibited cells versus Notch activated cells', DmD8 cells were split and one 1/2 pretreated with DFK-167 overnight. Untreated and DFK-treated cells were then exposed to EDTA and harvested after 30minutes. RNA was isolated using Trizol (Sigma) and reverse transcription was performed with Superscript III Reverse Transcriptase (Invitrogen) and oligo-dT primers (Sigma). Control and experimental samples were labeled with Cy3 or Cy5, mixed together and hybridized on Drosophila transcriptome long-oligonucleotide microarrays (FlyChip, FL002; http://www.flychip.org.uk/services/core/FL002/). Three biological replicates and dye swaps were performed for each experiment (5 arrays in total/experiment). Slides were scanned by Genepix 400B dual laser scanner (Axon) and spots were found and quantified by Dapple software. KC cells were obtained from Dr Martin Zeidler. Control versus Notch activated KC cells: mRNA was extracted from KC cells incubated in the absence or presence of EDTA to activate Notch (cells were harvested 30 minutes after addition of EDTA). RNA was isolated using TRIzol (Sigma) and reverse transcription was performed with Superscript III Reverse Transcriptase (Invitrogen) and oligo-dT primers (Sigma). Control and experimental samples were labeled with Cy3 or Cy5, mixed together and hybridized on Drosophila transcriptome long-oligonucleotide microarrays (FlyChip, FL002; http://www.flychip.org.uk/services/core/FL002/). Three biological replicates and dye swaps were performed (6 arrays in total/experiment). Slides were scanned by Genepix 400B dual laser scanner (Axon) and spots were found and quantified by Dapple software.