i want to create a stable cell line. problem is that the cells have already been immortalized with sv40 large t antigen. So i cannot use sv40 promoter since this would lead to episomal replication.
I need to use zeocin-resistance for selection, because my cells already have g418 resistance.
we have the zeocin-resistance-gene on another vector (pBudCE4.1), but under control of sv40 promoter.
I want to clone this resistance in pcDNA3.1
my question is: can i use the cmv early promoter for the zeocin resistance even though this is also the promoter for my insert. i have never seen a plasmid, where insert and resistance gene are under control of the same promoter..
thx for any answer.
Submit your paper to J Biol Methods today!
same promoter for insert and resistance??
1 reply to this topic
Posted 22 November 2010 - 02:40 AM
In case youre interested, i asked a professor. she said: people think that if 2 promoters of the same kind are on the plasmid they might hinder each other because they need the same factors. so the two promoters are in a competition which might lead to reduced expression of one or both of the products. To avoid such effects you prefere to have different promoters.