How do you pick a constitutively expressed promoter from a bacterial genome (to place upstream of an antibioticr gene in a knockout cassette) when you have the genome sequence?
I would like to design a linear (naked) DNA cassette to create a knock-out mutant by homologous recombination in a Bacteroides species.
I know that approx 500bp upstream and downstream of the target gene must be fused to a suitable antibiotic resistant cassette (Tet or Ery) but I am concerned that the current promoter region may not function in my bacterial species.
I would like to fuse a promoter from my bacteria to regulate expression of this resistance gene instead. The genome is sequenced but no genetic system exists for this bacteria.
Would anyone know if there are rules for picking a promoter for this purpose?
Does it have to be a housekeeping gene promoter that is always "on"?
Are there standard ones normally used, 16S/HSP/etc promoters?
Is it just trial and error with different ones?
Thanks in advance
Edited by Chris22, 30 March 2015 - 10:43 AM.