8 protocols
AccessionNameType
P-MTAB-27192
image_acquisition
Scanning was done using Agilent Technologies Scanner G2505B. Feature Extraction software was used for image analysis.
P-MTAB-27191
hybridization
825 ng of labeled sample (Cy5) and 825 ng of labeled reference sample (Cy3) were combined and a total of 1650 ng of combined and fragmented cRNA was used for hybridization. Combined experimental and reference samples were mixed with blocking agent, fragmentation- and hybridization buffer, and were hybridized to Agilent Mouse Genome Microarray 4x44K for 17 hours at 65 C
P-MTAB-27190
labeling
Labeling was done according to manufacturer's (Agilent) instructions. Primed cDNA was amplified and labeled using fluorescently labeled dNTP-nucleotides (Cy3 and Cy5) and T7 RNA Polymerase. Clean-up of labeled and amplified samples was performed by using Qiagen RNeasy mini spin columns. RNA Spike-In Kit was used to monitor the success of the labelingRNA Spike-In Kit was used to monitor the success of the labeling. The starting amount of total RNA was 1000 ng.
P-MTAB-27195
hybridization
Hybridization was done according to manufacturer's (Agilent) instructions. A total of 600 ng of fragmented cRNA was used for hybridization. Purified samples were mixed with blocking agent, fragmentation and hybridization buffer, and were hybridized to Agilent SurePrint G3 Mouse GE 8 x 60K Microarrays for 17 hours at 65 C.
P-MTAB-27193
nucleic_acid_extraction
*Sample set 2 (8x60K chips)*: Total RNA was isolated prostate two-month-old PAP-/- and PAP+/+ mice prostates (4 mice/group) with RNeasy Midi kit (Qiagen, Valencia, CA, USA). Prior to total RNA isolation prostates were stored in RNAlater, RNA stabilization reagent (Qiagen).
P-MTAB-27194
labeling
Labeling was done according to manufacturer's (Agilent) instructions. RNA Spike-In Kit was used to monitor the success of the labeling. The starting amount of total RNA was 200 ng. 200 ng of total RNA was primed using oligo-dT(T7) promoter primers and converted to cDNA with AffinityScript RNase Block (mix of enzymes, Agilent). Primed cDNA was amplified and labeled using fluorescently labeled dNTP-nucleotides (Cy3)and T7 RNA Polymerase. Clean-up of labeled and amplified samples was performed by using Qiagens RNeasy mini spin columns. RNA Spike-In Kit was used to monitor the success of the labeling .
P-MTAB-27188
specified_biomaterial_action
Two-month old PAP (ACPP) knockout mice were compared to wild type mice
P-MTAB-27189
nucleic_acid_extraction
*Sample set 1 (4x44K chips)*: Total RNA was isolated prostate two-month-old PAP-/- and PAP+/+ mice prostates (3 mice/group) with RNeasy Midi kit (Qiagen, Valencia, CA, USA). Prior to total RNA isolation prostates were stored in RNAlater, RNA stabilization reagent (Qiagen).