strain: Muta Mouse ID: 44 tissue: spleen gender: male
Treatment protocol
Adult male Muta™Mouse were administered Dibenzo[def,p]chrysene (DBC, CAS no. 191-30-0, purity ≥ 98%, Cambridge Isotopes, Tewksbury, MA) dissolved in highly refined olive oil (Sigma-Aldrich, Oakville, ON, Canada) at 0.005 ml/g body weight. The doses (2.0, 6.3, and 20.0 mg/kg-bw per day) were administered by oral gavage for three consecutive days. There were 15 animals in each dose group (5 harvested per each of 3 time points [i.e., 4, 24, and 72 h after the last exposure]), as well as in the vehicle control groups. Five mice from each dose group (and control group) were anesthetized using isoflurane gas and then euthanized via cervical dislocation followed by chest cavity opening 4, 24, or 72 h after the final dose.
Extracted molecule
total RNA
Extraction protocol
For all time points, the spleen was removed, flash-frozen in liquid nitrogen, and stored at -80°C.RNA was isolated, hybridized, and analyzed as described elsewhere (Malik, A. I., Rowan-Carroll, A., Williams, A., Lemieux, C. L., Long, A. S., Arlt, V. M., Phillips, D. H., White, P. A., and Yauk, C. L. (2013). Hepatic genotoxicity and toxicogenomic responses in MutaMouse males treated with dibenz[a,h]anthracene. Mutagenesis. 28, 543-554.)
Label
Cy5
Label protocol
200 ng of each sample was labelled with Cyanine 5-CTP (Cy5); universal reference total mouse RNA (Agilent Technologies) was labelled with Cyanine 3-CTP (Cy3).
Adult male Muta™Mouse were administered Dibenzo[def,p]chrysene (DBC, CAS no. 191-30-0, purity ≥ 98%, Cambridge Isotopes, Tewksbury, MA) dissolved in highly refined olive oil (Sigma-Aldrich, Oakville, ON, Canada) at 0.005 ml/g body weight. The doses (2.0, 6.3, and 20.0 mg/kg-bw per day) were administered by oral gavage for three consecutive days. There were 15 animals in each dose group (5 harvested per each of 3 time points [i.e., 4, 24, and 72 h after the last exposure]), as well as in the vehicle control groups. Five mice from each dose group (and control group) were anesthetized using isoflurane gas and then euthanized via cervical dislocation followed by chest cavity opening 4, 24, or 72 h after the final dose.
Extracted molecule
total RNA
Extraction protocol
For all time points, the spleen was removed, flash-frozen in liquid nitrogen, and stored at -80°C.RNA was isolated, hybridized, and analyzed as described elsewhere (Malik, A. I., Rowan-Carroll, A., Williams, A., Lemieux, C. L., Long, A. S., Arlt, V. M., Phillips, D. H., White, P. A., and Yauk, C. L. (2013). Hepatic genotoxicity and toxicogenomic responses in MutaMouse males treated with dibenz[a,h]anthracene. Mutagenesis. 28, 543-554.)
Label
Cy3
Label protocol
200 ng of each sample was labelled with Cyanine 5-CTP (Cy5); universal reference total mouse RNA (Agilent Technologies) was labelled with Cyanine 3-CTP (Cy3).
Hybridization protocol
Samples were hybridised to Agilent G3 Mouse GE 8x60K microarrays.
Scan protocol
Scanned on an Agilent G2565AA scanner.
Data processing
Images were quantified using Agilent Feature Extraction Software (version A.11.0.1.1). Non background subtracted median signal intensities were LOWESS normalized using the maanova library in R. Probes with technical replicates were averaged using the median.
