sampleID: 1 tissue: Part of lung age: 25 week gender: Male genotype/variation: transgenic mouse strain 40.6 exposure: 28 days, daily time post exposure: 3 days dosage: none
Extracted molecule
total RNA
Extraction protocol
RNA was isolated using the TRIzol reagent (Invitrogen Canada Inc., Burlington, ON, Canada), and further purified using RNeasy Mini Kits (Qiagen Inc., Mississauga, ON, Canada). RNA was quantified using UV spectrophotometer, and quality was confirmed using the Agilent 2100 Bioanalyzer and RNA 6000 NanoLab Chip Kit (Agilent Technologies Canada Inc., Mississauga, ON, Canada).
Label
Cy5
Label protocol
Individual 250 ng aliquots of RNA from each sample were amplified and labelled using the Low RNA Input Fluorescent Linear Amplification Kit (Agilent, (5184-3523)).
catalog#: 740100 vendor: Stratagene, La Jolla, CA, USA sample type: Total RNA derived from 11 mouse cell lines
Extracted molecule
total RNA
Extraction protocol
Unknown: Material was purchased as total RNA (universal mouse reference total RNA (Stratagene, CA, USA))
Label
Cy3
Label protocol
Individual 250 ng aliquots of RNA from each sample were amplified and labelled using the Low RNA Input Fluorescent Linear Amplification Kit (Agilent, (5184-3523)).
Hybridization protocol
Agilent Mouse G4846A Microarrays (containing approximately 40,000 probes) were hybridized with 250 ng Cy5-labelled liver RNA and 250 ng Cy3-labelled Universal Mouse Reference RNA (Stratagene, CA, USA), used as a common reference on all arrays. Arrays were incubated for 17 hours at 65 ºC in Agilent hybridization solution and washed according to manufacturer’s instructions.
Scan protocol
Arrays were scanned scanned on an Agilent G2505B scanner. Data were acquired using Agilent Feature Extraction software version 10.7.3.1
Data processing
The log2 of the ratio (sample(Cy5)/reference(Cy3)), using the median signal intensities were normalized using lowess using the transform.madata function in the maanova library in R.
