Laboratory evolution of new lactate transporter genes in a jen1 Δ mutant of S accharomyces cerevisiae and their identification as ADY 2 alleles by whole‐genome resequencing and transcriptome analysis

Laboratory evolution is a powerful approach in applied and fundamental yeast research, but complete elucidation of the molecular basis of evolved phenotypes remains a challenge. In this study, DNA microarray‐based transcriptome analysis and whole‐genome resequencing were used to investigate evolution of novel lactate transporters in S accharomyces cerevisiae that can replace J en1p, the only documented S . cerevisiae lactate transporter. To this end, a jen1 Δ mutant was evolved for growth on lactate in serial batch cultures. Two independent evolution experiments yielded growth on lactate as sole carbon source (0.14 and 0.18 h −1 , respectively). Transcriptome analysis did not provide leads, but whole‐genome resequencing showed different single‐nucleotide changes ( C 755 G / L eu219 V al and C 655 G / A la252 G ly) in the acetate transporter gene ADY 2 . Introduction of these ADY 2 alleles in a jen1 Δ ady2 Δ strain enabled growth on lactate (0.14 h −1 for A dy2p Leu219Val and 0.12 h −1 for A dy2p Ala252Gly ), demonstrating that these alleles of ADY 2 encode efficient lactate transporters. Depth of coverage of DNA sequencing, combined with karyotyping, gene deletions and diagnostic PCR , showed that an isochromosome III ( c . 475 kb) with two additional copies of ADY 2 C755G had been formed via crossover between retrotransposons YCLW Δ15 and YCRC Δ6 . The isochromosome formation shows how even short periods of selective pressure can cause substantial karyotype changes.