I would like to know if there is any genetic trick to repress the expression of a reporter on a plasmid but only before integration in the injected cell genome.
I'm trying to get rid of the expression from the naked plasmid injected in cells, but make it visible only when it is inserted.
Thanks for your answer!
Hi Thomas,
I'm not sure if you're talking about eukaryotes or prokaryotes. I only have genetic experience with bacteria, so I'll answer based on that and maybe it can help you.
1. Engineer the reporter without a promoter, and then integrate it into the genome immediately downstream of a native promoter. The reporter gene can then tell you about native signals that control gene expression.
2. Use a very tightly repressible promoter (i.e. rhamnose promoter) and only add inducer once the clones are single-colony purified and you have checked for the integration event.
Hope this helps!
Andrew
Dear Andrew,
Thanks for your answer, but I was talking about eucaryotes specifically. Sorry for the confusion.
I think you can use tetracycline induced systems to control the expression of your heterologous genes.
Please refer to:
Article How to Choose the Right Inducible Gene Expression System for...
I'm not sure, but if the integration of your gene is precisely under your control, maybe you can design your genetic construct in a way that would form a complete promoter sequence only upon integration to the genome.
Dear Mohammed, thanks for your answer. But the system you proposed won’t work. Let me rephrase the question maybe: I’m looking for a genetic trick so that if we inject a plasmid in a eucaryote cell, we would have a trancription of a transgene from the genomic integrated plasmid only, and not from episomal copies.
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