PM was induced with the syngeneic colon adenocarcinoma CC531 cells (purchased at CLS Cell Lines Service GmbH, Eppelheim, Germany, product number 500,387). The cells were cultured in RMPI 1640 medium (Thermo Fisher Scientific) supplemented with 10% fetal bovine serum (Sigma-Aldrich, Zwijndrecht, the Netherlands) in a 5% CO2/air incubator at 37 °C.
CC531 cells were transduced with a neomycin-resistant firefly luciferase gene, as previously described [28 (link)]. Briefly, lentiviral particles were produced by transfecting HEK 293FT with the vectors pLenti PGK V5 luciferase (Addgene plasmid #21,471), pVSV-G, and pCMVd8.74. CC531 cells were transducted with these lentiviral particles, resulting in CC531-Luc cells. Selection with neomycin 2 mg/ml (Sigma-Aldrich geneticin G418, Zwijndrecht, the Netherlands) began 48 h after the transduction process to isolate the resistant colonies positively tested for luciferase activity. After at least three weeks of continuous selection, cells were used in experiments. Before injecting into the animals, the cells were tested for the absence of mycoplasma and rat-specific viral pathogens.
CC531 cells were transduced with a neomycin-resistant firefly luciferase gene, as previously described [28 (link)]. Briefly, lentiviral particles were produced by transfecting HEK 293FT with the vectors pLenti PGK V5 luciferase (Addgene plasmid #21,471), pVSV-G, and pCMVd8.74. CC531 cells were transducted with these lentiviral particles, resulting in CC531-Luc cells. Selection with neomycin 2 mg/ml (Sigma-Aldrich geneticin G418, Zwijndrecht, the Netherlands) began 48 h after the transduction process to isolate the resistant colonies positively tested for luciferase activity. After at least three weeks of continuous selection, cells were used in experiments. Before injecting into the animals, the cells were tested for the absence of mycoplasma and rat-specific viral pathogens.
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