description: Protocols: 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 (Kühner, Switzerland) at 37°C, 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 µg/mL final concentration) in cultivation medium for four hours before cells were harvested. Cells were washed twice in PBS and stored at -80°C 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 manufacturer's protocol.
attributes.tag: ENA-FIRST-PUBLIC
attributes.value: 2022-08-09
attributes.tag: ENA-LAST-UPDATE
attributes.value: 2022-08-09
attributes.tag: External Id
attributes.value: SAMEA11293139
attributes.tag: INSDC center alias
attributes.value: The University of Manchester
attributes.tag: INSDC center name
attributes.value: the university of manchester
attributes.tag: INSDC first public
attributes.value: 2022-08-09T00:23:53Z
attributes.tag: INSDC last update
attributes.value: 2022-08-09T00:23:53Z
attributes.tag: INSDC status
attributes.value: public
attributes.tag: Submitter Id
attributes.value: E-MTAB-11173:Sample 1
attributes.tag: broker name
attributes.value: ArrayExpress
attributes.tag: cell_line
attributes.value: CHO-K1
attributes.tag: common name
attributes.value: Chinese hamster
attributes.tag: developmental stage
attributes.value: adult
attributes.tag: genotype
attributes.value: 2G12
attributes.tag: organism part
attributes.value: ovary
attributes.tag: sample name
attributes.value: E-MTAB-11173:Sample 1