Hi everyone. I know that for an integrative plasmid you must add homology sequences to an specific locus where you want the integration, but i have doubts about regions that are usually prefered for IYp. In addition i want to know how to design that. I mean if i already have my construct with my gene of interest, should i add that homology sequences at both ends of my construct, and what should be the lenght of the homology region?
Hy Sebastian, I can suggest to link your DNA-fragment in the middle of LYS or URA genes carried on specific plasmid, and proceed with ligation having digested both with the appropriated enzymes. Pay attention to choose the right enzymes. Good luck
If your gene of interest is a yeast gene then it will serve as the region of homology. But it not, then yes you would need to add a segment of DNA that can serve as a region of homology. Additionally you need to have an appropriate selectable marker for the yeast strain you are using. Depending upon the yeast strain it might be possible that the selectable marker can also be your region of homology but generally most labs use deletion strains of the selectable marker (like URA3) in which case obviously there would no longer be homology.
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