I cloned 2 genes of my interest into the pBI-4 vector, so that I could activate the expression of both genes at the same time. However, the construct is quite leaky, so that I get expression of proteins without adding Doxycyclin. I want to generate a stable cell line, therefore I would need a tightly regulated promoter, since the constitutive overexpression of these proteins is cytotoxic.
vector with the TRE-tight response element and minimal promoter are probably to best to solve your problem of leakyness, but I dont know if there is a pTRE-tight plasmid with bi-directional expression.
Dear Elvira,
leaking using cell culture with regular serum comes from the presence of tetracyclin antibiotic into it. You can either remove tetracyclin by removing lipids or buy antibiotic-free serum.
Good luck ,
Edouard PROST, PhD
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