E-mail From NEB on Dec 9th

2015-01-13

azim58 - E-mail From NEB on Dec 9th

Dear Kurt ,

Thank you for contacting NEB Technical Support. Your question and answer
are below.

Your question:
Oh actually it sounds like you may have exactly what I want! I want to
express an scFv (single chain Fragment variable) protein as a fusion with
the p3 M13 protein. An scFv gene is only about 750 bp and is like a mini
antibody containing only the heavy chain variable region, light chain
variable region, and a linker connecting the two. I think I can just
insert my scFv gene sequence instead of a sequence for a peptide if I am
not mistaken. Does your vector contain a SfiI site and NotI site that
would allow me to clone my peptide or scFv sequence into the vector?

Could I get a link to your system(s) applicable for this? Thanks for any
information you can offer me!

Our answer:
We have anecdotal evidence that large peptides may be displayed attached
to pIII but we generally say 50 amino acids is the max. We do know some
peptides/fragments display better than others so your 750 bp fragment is
worth a shot. Unfortunately, to keep the pIII leader sequence intact, we
did not design multiple restriction sites at the pIII insertion point.
You may be able engineer a couple others in there but we exclusively use
KpnI and EagI for cloning. The phage display vector is called M13KE and
the RF DNA is available in #E8101S PhD Phage Display Cloning System,

http://www.neb.com/nebecomm/products/productE8101.asp

Regards,

Beth

azim58wiki: