Hello everyone,
I wondered what's the advantage to use IPTG in place of Doxycycline for gene expression modulation in vivo ?
I already eared that DOX can become be toxic for mice in case of long term treatment, while IPTG doesn't affect the clinical signs or the general health of mice.
Do you have an idea about that?
Thanks in advance
And Merry Christmas!
I am not aware of using IPTG for gene induction in mice. People express lacZ in mice as a marker for gene expression, but it is usually under the control of a different promoter. IPTG induction is used to activate the lac operon in E. Coli. Doxycycline is used in the Tet-on or the Tet-off system. These are engineered using the tetracycline transactivators that can be either activated by Dox (Tet-on) or repressed by Dox (Tet-off).
Hi,
I would vote for IPTG over Dox. Personally I have used Dox system which is killing my 293T cells at above 5ug/ml concentration. If you have a system already installed, using Lac operon (IPTG as an inducer) please try it and see the difference concerning cell death.
there is a report as well on TCNA toxicity:http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0114457
Anand
Thanks a lot for the answers!
In fact, I'm using IPTG for expressing an shRNA (displayed in a pRL TK LacO plasmid) and it works perfectly. But people asked me why I din't use Dox, and they never eared about using IPTG in vivo, but only in batecrial procedures... So I asked the suggestion of users about that, and it validates our decision to use the IPTG system, Thank you!
Yes you are right, IPTG is something synonymous to use in Bacteria. you may get questions from everyone. To show your system is better, I would simply make cell viability/proliferation data in presence of IPTG and Dox in a diffenent concentration. I assure you at 10ug/ml Dox, you will see only dead cells on the plate.
Good luck, please share me informaton on your IPTG system (a paper or company web link)
thanks,anand
The one that we used is a Mission shRNA Vector from sigma (pLKO-puro-IPTG-3xLacO) and it treat my cells (GBM cells) with 1mM routinely in culture and in vivo I perform injection IP at a concentration of 20mM (200uL injection) but people also put it in drinking water at a concentration of 10mM. The only disadvantage is the sulfur smell… But it works really well for protein depletion!
Depending on whether you want to re-express a target protein or knock it down you might wanna use Dox only or IPTG/Dox respectively.
You cannot put whatever you want in your LacO plasmid? Here we put a sequence for shRNA but we could have put a sequence coding for a protein for example, also using the IPTG inducible system. But maybe I'm wrong?