(A) The plasmid used in this study. Each glycolytic gene was cloned into a 2-µm-based multicopy plasmid (pTOW40836) and expressed from the TDH3 promoter (TDH3pro) (with the exception of PYK1 which was expressed from the PYK1 promoter [PYK1pro], represented in the figure as (p)). In +leucine conditions, the copy number of the plasmid is relatively low (~30). In –leucine conditions, the copy number goes up to 150 copies per cell due to the bias of leu2-89 (Moriya et al., 2012). Here, we designate these conditions low- and high-copy conditions, respectively. (B and C) Maximum growth rate of yeast cells harboring the plasmid overexpressing each glycolytic protein in the indicated growth conditions. The unit is min−1 × 10−4. (D and E) Copy number of the plasmid overexpressing each glycolytic protein in the indicated growth conditions. The unit is copy number per haploid genome. The error bars shows the standard deviation of at least three independent biological measurements.