I want to know, when you are working with an environmental isolate, how authentic it is to use plasmid base system for homologous expressions as host recombination machinery might integrate as its cultured subsistently? Is chromosome integration a must if plasmid replicon is stable?
Your question is a little vague in that I am not sure what strains and plasmids you are using. But assuming that the plasmid is naturally maintained by the strain integration should not be a problem. I am not sure what you mean by homologous expression - I can go two ways either you mean homologous recombination or homogeneous expression. The first would fit with recombination - this would be an issue if the plasmid is not able to replicate in the host, and so would be essential for the maintenance of the resistance marker (on the plasmid) as it would be the only way that the marker would be inherited efficiently. The second option is more problematic and would depend on how the gene of interest is expressed - my bet would be that the expression would not be the same for all cells in a population.
Basically to help I need more information and careful use of the terminology.
Thanks Richard, for your constructive comment to make the question more apt to answer, As you have very appropriately stated about the terms Homologous recombination and Homogeneous expression. Making thing more clear, I am working on Azoreductase in Enterococcus sp. and I want to increase the basal AzoA level in the same host by using pbacA ( bacitracin resistance gene promoter) to drive constitutive expression by plasmid borne system.
SO I am questioning that since, Enterococcus sp. strain is natural strain having recombination ability and azoA gene cassette is the homologous region available here by single cross-over event might lead to integration of the plasmid on chromosome and cells will be still remain resistant since the linearized chromosome integrated plasmid segment will carry the antibiotic resistance gene. Thus I am regularly checking for presence of plasmid in the cells but this question still remains to justify my work.
Dear Rahtod
Please find the attached file. Hope to be benefit
https://en.wikipedia.org/wiki/Regulation_of_gene_expression
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