Protocol Online logo
Top : New Forum Archives (2009-): : Molecular Cloning

Cloning a fusion protein, do I remove the ATG codon. - (Aug/15/2014 )

Hi all,

I have a dilemma if anyone was kind and knowledgeable enough.

I am cloning a N-terminal fusion protein, and I am designing my primers so that I have to remove the ATG codon of my gene of interest.

 

My question --> is it appropriate to remove the ATG from your protein and rely on the transcription of you fused protein within the vector?

 

I am little stumped on why you have to remove the ATG in the first place?

 

Would appreciate some ideas.

Cheers

 

Yours Sincerely Liam Tegg

-Teapot_11-

You don't need to, but if you are using a short tag you run the risk that the RNA pol will skip over the tag and start transcription at the internal ATG. To get around this, you can add a Kozak (or Shine-Delgarno, depending on application) sequence at the ATG for the tag.

-bob1-