3 replies to this topic

[saimasbs]

i m working on protein expression . i hav only  available option to cut my gene with Nde1 or Nco1 to express it in pet system but these enzymes r cutter of my gene also. plz guide me how can  i get pure protein of my gene .

[bob1]

PCR out the gene using primers with appropriate restriction sites, then clone the PCR product into your vector of choice.

[saimasbs]

thanks bob1 for replying.i hav cloned my gene in PTZ57R/T vector.now i hav to express and purify protein .my gene size is about 3kb and i hav introduced ECORI AND HINDIII sites in it for clonning it in pet21a for expression. but there will be some extra protein with my gene product so how can i will i purify it?

[bob1]

How will there be extra protein?  Your restriction sites should be outside the start and stop codons...