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. 2020 Dec 2;117(50):31789–31799. doi: 10.1073/pnas.2013968117

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Fig. 3. — Determining the limiting steps in the peroxisomal pathway. Combinatorial overexpression of the peroxisomal (R)-(+)-limonene pathway genes in strain PERLim09 to determine the limiting steps. (A) Strain PERLim09 was transformed with high copy-number episomal vectors overexpressing ClLimS and/or Erg20p(N127W) (or the corresponding empty vector as control) and the effect of the up-regulation of the corresponding enzymes on (R)-(+)-limonene production was evaluated. (B) PERLim09 was transformed with high copy-number episomal vectors overexpressing the pathway genes in pairs (IDI1 with ERG19, ERG8 with ERG12, and EfmvaS with EfmvaE) under conditions where ClLimS and Erg20p(N127W) were also overexpressed. (C) The contribution of individual overexpression of Erg12p and Erg8p was evaluated by transforming PERLim09 with high copy-number episomal vectors under conditions where ClLimS and Erg20p(N127W) were also overexpressed. Error bars correspond to the SD of the mean (n = 3, corresponding to three biological replicates).