2014-08-11 Progress Report

Choosing a Vector

The second component of the vaccine is the vector (plasmid backbone). The one major criterion here is that the vector must be strongly and constitutively expressed in mammals.

Ideally, the vector is specifically designed for protein expression in mammals: i.e., it has a leader (signal) peptide, a Kozak sequence, a transcription termination signal, and optimizations to improve expression, such as enhancers.

Many vectors include myc epitopes and/or polyhistidine tags to enable you to recover the recombinant protein. With a DNA vaccine, we are not attempting to harvest the recombinant protein so this stuff is unnecessary, and may even be detrimental.

I first looked around for a suitable BioBrick vector, but I could not find one. I found a reference to addgene.org that seemed promising, but the plasmids I found there were limited to non-commercial use.

Although I'd prefer not to use a commercial vector, there are a few that seemed promising.

• Invitrogen/Life Technologies pSecTag2
• clonTech pIRES: The main advantage of this vector is that it can express two proteins from one CMV promoter thanks to an IRES
• NEB pMCP