We would like to label E. coli HB101 with GFP and also have it carry a plasmid (pBR322 ori) with another gene of interest. Since it is a RecA- strain, we were looking at the pOSIP plasmid (St-Pierre et al. 2013) method of chromosomal GFP integration. But since HB101 is not a T7 (DE3)strain, what promoter would be the best to use for constitutive expression of GFP? Our end goal is to be able to co-localize the GFP E. coli with other fluorescent antibody tagged cellular markers (after PFA/methanol fixation).
The em7 promoter is used pretty frequently for strong constitutive expression in E. coli.
Another option would actually be the λPL or λPR promoter since those are strong and constitutive in the absence of the λcl repressor, but only go for that if you know the repressor will be lost after the integration steps, I'm not really sure how the pOSIP system works exactly.
The other approach to consider is to use a second plasmid with different compatibility groups from pBR322. This might be easier for you to engineer.
We decided to do two different ways. We used the EM7 constitutive promoter with a superfolded GFP on a p15A origin plasmid and we also integrated the fluorophore with promoter into the chromosome using the clonetegration (pOSIP) method.
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