I want to C-terminally His tag a protein from Caulobacter (a gram-negative
bacteria), and I am thinking about using pET-28a vector from Novagen. But
this vector carries both an N-terminal His tag and an C-terminal one. If I
clone my gene in frame with the C-terminal His tag, do I need to clone the
ribosome binding site of that gene from the Caulobacter (but I want to
express it in E.coli)?? The order of the sequence in that vector is: rbs--N
terminal his tag--restriction site--C termianl his tag
And do I need to remove the N-terminal his tag??
If I need to clone the rbs from the Caulobacter, is there any software or
tool that I can use to search for the ribosome binding site of a gene?? I
don't know the exact sequence of rbs in Caulobacter, and I think it's
different from E.coli
Help will be much appreciated!! Thanks!!