Deletion of a KU80 homolog enhances homologous recombination in the thermotolerant yeast Kluyveromyces marxianus

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Abstract
Targeted gene replacement in the thermotolerant yeast Kluyveromyces marxianus KCTC 17555 has been hampered by its propensity to non-homologous end joining (NHEJ). To enhance homologous recombination (HR) by blocking NHEJ, we identified and disrupted the K. marxianus KU80 gene. The ku80 deletion mutant strain (Kmku80∆) of K. marxianus KCTC 17555 did not show apparent growth defects under several conditions with the exception of exposure to tunicamycin. The targeted disruption of the three model genes, KmLEU2, KmPDC1, and KmPDC5, was increased by 13–70 % in Kmku80∆, although the efficiency was greatly affected by the length of the homologous flanking fragments. In contrast, the double HR frequency was 0–13.7 % in the wild-type strain even with flanking fragments 1 kb long. Therefore, Kmku80∆ promises to be a useful recipient strain for targeted gene manipulation.

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