How can I change a promoter of a plasmid? - (Sep/03/2005 )
I want to change the promoter of a yeast plamid to another one, does anyone know how to change the promoter?
Since there may be a RBS(Ribosome Binding Site) following the promoter, is RBS sequence specific and how to design it?
There's a couple ways:
1. The most efficient way is to use restriction digest to lift out the existing promoter. Amplify your new promoter with sticky ends compatible with the REs used to remove the plasmid promoter and ligate in.
2. Depending on th size of your plasmid you can amplify the plasmid without the promoter including RE sites you choose in the oligos. Then you can take your amplified promoter with sticky ends to the plasmid sticky and ligate together.
As far as the RBS, I'm not positive, but it may be specific to the gene being expressed, but not specific to the promoter.
hope this helps...