title: Genotyping of Scer harboring FCY1 promoter mutations after bulk competition, using a shorter sequencing fragment
design: S. cerevisiae yeasts transformed with a pool of FCY1 promoters each harboring a single mutations were grown in SC media with or without 5-FC [12 uM] for 20 doublings, with intermediate dilution steps to maintain exponential growth (T0: start; T1: first dilution; T2: second dilution; T3: end). Each pool of mutants was generated in four biological replicates, each used to inoculate two technical replicates. Each technical replicate was also repeated with the addition previously characterized mutations in the FCY1 gene (CDS) as controls. From all collected cell pellets, genomic DNA was extracted and two rounds of PCR were used to specifically amplify the 154 bp mutagenized region of the promoter (F3-F4). For the samples with added CDS mutations, additional PCRs were perform to amplify a short segment of FCY1, specifically in mutants from the control CDS set.
identifiers.namespace: SUB15610361
identifiers.id: F3F4_and_CDS_150