Selection strategies for the establishment of recombinant Chinese hamster ovary cell line with dihydrofolate reductase-mediated gene amplification.
To evaluate the efficacy of selection strategies for recombinant Chinese hamster ovary (rCHO) clones undergone with dihydrofolate reductase-mediated gene amplification, rCHO cell lines producing a chimeric antibody were established using two strategies, one based on individual clones and the other based on cell pools. In a selection based on individual clones, cell cloning by limiting dilution method was performed twice, once after a round of selection of parental cell clones and once after obtaining high-producer clones. Thirty parental clones selected from 300 parental clones were cultivated independently throughout the gene amplification procedure. Using this labor-intensive strategy, it took approximately 17 weeks to obtain high-producing clones such as CS11-8 and CS18-3 clones. A selection based on cell pools, in which cell cloning was performed once at the final selection stage, required less effort and time to amplify large numbers of individual parental clones within the pool. However, high-producing clones were lost during the amplification procedure. The antibody expression level of high-producing clones such as PS7-2 and PS7-32 chosen on the basis of cell pools was less than one third of that of CS11-8 and CS18-3 clones. Taken together, a selection strategy based on individual clones is favored for establishment of high-producing rCHO clones because it is more efficient to perform cell cloning at the initial selection stage of parental cell clones.
Jun
Seung Chul
SC
Department of Biological Sciences, Korea Advanced Institute of Science and Technology, Yusong-Gu, Daejon.
Kim
Min Soo
MS
Baik
Jong Youn
JY
Hwang
Sun Ok
SO
Lee
Gyun Min
GM
D000818:Animals; D016466:CHO Cells; D018929:Cell Culture Techniques / Q000379:methods*; D002999:Clone Cells / Q000166:cytology / Q000187:drug effects / Q000378:metabolism; D006224:Cricetinae; D003412:Cricetulus; D005784:Gene Amplification; D008727:Methotrexate / Q000494:pharmacology; D021141:Nucleic Acid Amplification Techniques; D012641:Selection, Genetic; D013762:Tetrahydrofolate Dehydrogenase / Q000235:genetics / Q000378:metabolism*
D013762:Tetrahydrofolate Dehydrogenase; D008727:Methotrexate