Hi everybody,
I want to co transform 2 genes that are in different plasmids into one Cell, they are both in a same plasmid, so if I do they are incompatible (they have pBR322 ori)
So I need to use another plasmid for one of the genes (pA15 ori), to don't waste time in ordering pCyC plasmid, I want to use pLySs (that have pA15 ori).
However, I read that this plasmid in not for cloning but for expression of T7 RNA terminator.
Can I still use it to clone my gene?
Thanks for your help.
Hello Oualid,
pLysS and pLysE express the T7 lysozyme, which quenches the residual activity of T7 RNA polymerase in the absence of inducer. They are not designed for recombinant protein production. If you want to co-express two genes, I suggest that you clone them in a vector like pETDuet, which provides two independent multiple cloning sites under the control of two T7 promoters
Cher Pierre Béguin,
Merci beaucoup pour votre réponse.
Effectively, pETDuet is an option but i am lacking of time so that's why i ask for pLySs since i have it in the lab.
Thanks again.
Cordially
There is no problem for you to clone your gene into pLySs vector but you probabaly hardly to detect your target protein expression with this vector since it does not have a inducible strong promoter and usually has a very low level of T7 lysozyme expression in E. coli strains.
As others have rightly advised, pETDUet is your best bet. http://www.helmholtz-muenchen.de/fileadmin/PEPF/pET_vectors/pETDuet-1_map.pdf . pLySs is not used to expressed target genes.
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