lentiviral systems for overexpression: clone in full cDNA or ORF? - (Aug/20/2008 )
I use lentiviral packaging system from Open Biosystems and am about to make some new constructs for gene overexpression. While ordering cDNA clones from Origene, a tech support specialist told me that if I am going to use lentiviral systems I am better of cloning in ORFs rather than the full lengths cDNAs with 5' and 3' regions because polyA tail on the clone may inhibit lentiviral packaging. I have never come across this bit of information and can not find any references on-line to support or disprove it. I am just a numbnut lentiviral system user and know very little about their biology. Does anyone have any input on this topic?
Thank you very much,
Christine
Never heard of that neither, but still maybe possible.
I would go with the ORF, unless you have evidence the UTR regions could play a role in the protein's expression in some way (mRNA stability, traductability, etc)
I would go with the ORF, unless you have evidence the UTR regions could play a role in the protein's expression in some way (mRNA stability, traductability, etc)
Thank you Madrius!
I am putting in ORFs and will try one construct with and without UTRs just in case.
christine.
Some proteins are expressed better with the UTR's. That's protein dependent. Also poly A is not used in lentiviral vectors that we have or our collaborators.