[Microbiology] Suggestion for primer designing for expression

[shrinivas Dengeti]

Hi friends..

I am working with pET vector system. for expression of the enzme in
E.coli BL21.
I have to design primer with Nco1 site in forward primer.
so can anybody will suggest me about the placing of ATG site of
ORF...whether it should be in the primer's R.E site or else again i
have to add in the primer aftre the R.E site.


waiting for suggetion from scientific community