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Tet-responsive cell line - cells cannot become confluent (Aug/10/2004 )

Hello, thank you for reading my post.

I am developing a tet-responsive line from a cell type that starts to differentiate when the cells become confluent. Therefore, during the selection process, I will either have to continue splitting the cells and pool clones, or I will have to let colonies form and hope they do not change very much. Does anyone have any suggestions as to any other options? I was thinking about limited dilution to try to get single clones, but I will have to screen so many colonies because they will be repetetive, if that makes any sense. Meaning, many of them will have come from the same colony in the first place. I very much appreciate any help you can give to me.

Thank you,


Sorry, I'm a little vague on how long it take for these cells to differentiate. Are you saying if you put one cell in a well and let it double say 5-8 times, that will put enough pressure on it to start differentiation?

If so, even limited dilution will no work. Am I missing something?