Each dot is a transfected host cell carrying a random number of
integrated copies of the product gene. Copies inserted into active,
transcriptionally-favourable chromatin ("hot spots") produce more
protein per cell per day; copies landing in silenced regions produce
little. Every passage applies a selection agent that kills the
lowest producers, then the survivors are expanded back up to a full
population — with a little random drift added, since epigenetic
silencing continues even after selection.
titer(clone) ~ f(copy_number, integration_site)
survivors = top (1 − stringency) fraction by titer
stability(passage) = mean_titer(passage) / mean_titer(passage 0)
- Selection stringency — fraction of the population killed each passage by the selection agent (e.g. methotrexate); higher stringency enriches producers faster but shrinks diversity.
- Vector copy number — average gene copies per cell; more copies raise ceiling titer but increase instability risk.
- Mutation drift — random titer change per passage from ongoing epigenetic silencing/genetic drift.
- Targeted integration — site-specific insertion (e.g. via recombinase) into a known safe-harbour locus, which narrows the titer spread compared with random integration.
This is the core logic behind real CHO cell line development: dozens
to thousands of clones are screened over several passages until one
stable, high-titer "master cell bank" candidate is chosen.