Deletion of POX3 gene from Yarrowia lipolytica

The POX3 gene could encode acyl-coenzyme A oxidase(acyl-CoA)in Yarrowia lipolytica and get involved in biosynthesis of γ-decalactone. POX3 knockout was mediated with a loxP-KanMX-loxP cassette centered in a PCR fragment employing 80bp primer pairs in which 60 nucleotides fully match the POX3 open-reading frame. After transformed the linear disruption cassette into Yarrowia lipolytica cells,selected transformants were checked by PCR for correct integration and deletion of the target sequence. The KanMX marker was efficiently removed by transforming pSH65 plasmid and inducing the Cre expression. pSH65 was then lost after continually incubated in YPD. At last,the knockout mutant of POX3 gene was generated. The research provides a basis for combination knockouts of other genes that inhibit γ-decalactone production.