Protocols: The RNA-seq experiment within this study was designed to identify radiation-responsive and IRF7-responsive genes in a prostate cancer cell-line, PC3. To do so we generated stable IRF7 knockdown derivatives using shRNA (referred to as 'IRF7' in the files) and scrambled controls (referred to as 'scr'). We subjected 5 million cells per condition to 1,2 or 3 doses of irradiation or mock irradiation with 24 hour or 72 hour recovery periods interspersed (annotated for example as '24-1', one dose with a 24-hour recovery period). At 24 hours or 72 hours after the final treatment we harvested cells (see below) The PC3 cell-line was grown in RPMI containing 10% FBS and penicillin and streptomycin. We subjected 5 million cells per condition to 1,2 or 3 doses of irradiation or mock irradiation with 24 hour or 72 hour recovery periods interspersed (annotated for example as '24-1', one dose with a 24-hour recovery period). Total RNA extraction was performed on adherent cells at the indicated timepoints (see above) using Trizol. To assess the yield and integrity of mRNA, 2ul of each sample was taken for analysis on a Fragment Analyzer Automated CE System (Advanced Analytical) and smear analysis run using a DNF-473 Standard Sensitivity NGS Fragment Analysis Kit (Advanced Analytical). Total RNA (1ug) was processed to first-stranded cDNA using the TruSeq Stranded mRNA Library Prep Kit (Illumina), following the manufacturer's instructions for the low-throughput protocol. First-stranded cDNA samples were processed to barcoded cDNA libraries using a Diagenode IP-Star, according to the manufacturer's instructions for Illumina TruSeq library prep. Resulting libraries were PCR-amplified to incorporate unique indices and cleanup performed using AMPure XP Beads (Becman Coulter) according to Truseq LT protocol.
ENA-FIRST-PUBLIC
2021-05-03T00:21:56Z
ENA-LAST-UPDATE
2020-07-01T16:55:42Z
External Id
SAMEA7032939
INSDC center name
Centre for Cancer Research and Cell Biology, Queen's University Belfast, United Kingdom, BT9 7AE
INSDC first public
2021-05-03T00:21:56Z
INSDC last update
2020-07-01T16:55:42Z
INSDC status
public
Submitter Id
E-MTAB-9286:AR8
broker name
ArrayExpress
cell line
PC3
common name
human
developmental stage
adult
disease
adenocarcinoma
metastatic site
bone
organism part
prostate
sample name
E-MTAB-9286:AR8
scientific_name
Homo sapiens