Name: Sample 7_p
Instrument: Illumina HiSeq 4000
Strategy: RNA-Seq
Source: TRANSCRIPTOMIC
Selection: PolyA
Layout: PAIRED
Construction protocol: The development of the isogenic, IgG-producing CHO cell lines was previously described (PMID: 31350616). Recombinant CHO-K1 expressing 353/11 and 2G12 (for short 353/11 and 2G12) were cultivated in triplicate in chemically defined CD-CHO medium (Gibco, #10743029), supplemented with L-glutamine, phenol red and ganciclovir. To ensure identical cultivation conditions and provide best conditions for a nearly identical physiological status of the cells we chose a semi-perfusion process mode in 50 ml vent cap spin tubes in an ISF-X shaker (Khner, Switzerland) at 37C, 80% humidity, 5% CO2 and 220 rpm. The seeding density was 5x106 cells per mL. Every 24 hours the cells in culture were counted, mAb titers as well as key metabolites were analyzed and the complete medium was exchanged. Cell pellets for transcriptomics analysis were taken on day six where both cell lines have reached maximum cell density and viability was continuously high. The sampling point was chosen four hours after medium exchange to avoid effects due to medium limitations. To induce ER stress, two additional 353/11 IgG cultures were treated with tunicamycin (TM, 1 g/mL final concentration) in cultivation medium for four hours before cells were harvested. Cells were washed twice in PBS and stored at -80C before nucleic acid isolation using TRIzol. Biological replicates (BR) were generated from different cell culture experiments while technical replicates (TR) were generated from the same culture and sampling point with multiple cell pellets Illumina TruSeq Stranded mRNA kit, according to the manufacturers protocol.