Dear collegues,
I want to produce double mutants by disrupting genes in Sachharomyces. Previously I have done gene disruption using the KanMx cassette. I was thinking to produce the second mutation utilizing a auxotrophic marker like URA3 or HIS3 or LEU2 for easier selection. But I am having difficulty in finding a plasmid source from where I can procure such cassette in order to perform the deletion/disruption. Please suggest.
You could order plasmids from Addgene (https://www.addgene.org/). Or just try requesting plasmids from other yeast labs. They should have it.
https://www.addgene.org/
Hi Rajaneesh
I will order the plasmids from Addgene but for that i need the info regarding which plasmid to order. I am asking around in other labs who work with yeast.
Thanks.
Hi Shakri,
I have ordered all of my S. cerevisiae vectors via the National BioResource project:
http://yeast.lab.nig.ac.jp/nig/index_en.html
They are much cheaper than addgene as it is a repository.
I routinely use pRS315 which has the LEU2 marker and pRS313 which is HIS3.
Regards,
Kerry
No problem. If you need any more info on cerevisiae plasmids or transformations just let me know.
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