|Application:||Gene expression microarray analysis|
|Number of samples:||6|
|Release date:||Dec 21 2005|
|Last update date:||Dec 6 2012|
|Diseases:||Neural Tube Defects, Werner Syndrome|
|Dataset link||Differential expression of genes in cells mutant for Wrn and/or PARP-1 compared to wild type cells|
Microarray analyses were performed on cell cultures at the second passage (10 population doublings). For each genotype, asynchronously dividing cells derived from three healthy 15.5 day embryos from one litter were pooled at the second passage and cytoplasmic RNA was extracted. Cytoplasmic RNA was used in these experiments to avoid contamination with heterogeneous nuclear RNA and genomic DNA. (Note that DNAse treatment was also applied to all samples). This pool of RNA was labeled sample number 1 for each genotype. Pooling of embryonic cells was performed to minimize the effect of inter-individual biological differences. A second pool of embryonic cells was also created from a separate dam (second litter) for each genotype (called samples number 2). This strategy allowed obtaining samples in duplicate for each genotype. The cRNA from wild type cells were synthesized with Cy-5 labeled nucleotides and cRNAs from Wrn mutant, PARP-1 null, and PARP-1 null/Wrn mutant cells were synthesized with Cy-3 labeled nucleotides. Hybridization was performed on Mouse Agilent 60-mer Oligo Microarray chips by mixing wild type labeled cRNA (baseline expression levels) with either Wrn mutant, PARP-1 null, or PARP-1 null/Wrn mutant cRNA. Hybridization experiments were done in duplicates.