Hello people. I have a question. Can I amplify the gene with a dominant maker if my gene already has an auxotrophic marker (already made strain) next to it? Will that auxotrophic marker cause any issue in the process and then can I use the dominant marker for the screening process and not the auxotrophic one because the strain in which I am trying to integrate this gene already has the auxotrophic marker for scoring the process of gene silencing. Please provide valuable inputs and the methodology I can use to proceed with the same. Thank you.
Does your plasmid have any antibiotic resistance markers? Also what species are you using?
The plasmid which I want to use is of pYM series given in Jankee etal,2004 paper. Different plasmids are there like for hygromycin, kanamycin and nourseothricin. I want to use any one of these dominant markers and I am using budding yeast for the process.
Assuming the promoter for the antibiotic selection markers will work in your yeast then you should have no problems.
But I have one more question . The gene has one more auxotrophic marker inserted next to it , so the tagging primers which I'll be using for the purpose. Shall one of them should be homologous to the marker or both of them should be homologous to my gene of interest? I want to amplify my product that is why asking
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