The first twenty data entries include genomic sequences derived from 20 diploid strains that have been selected to have recombination between the centromeres of the homologs of chromosome III. The diploid was constructed by mating of two sequence-diverged haploids (W303-1A and YJM789) and is heterozygous for approximately 55,000 single-nucleotide polymorphisms (Sui et al., 2020 PNAS 117: 28191-28200). Whole-genome sequencing of yeast strains was performed on the Illumina HiSeq 2500 sequencer using a paired-end indexing protocol. Mapping of the sequencing reads and point mutation calling were performed as described in our previous study (Qi et al., 2023 PLoS Genetics 19: e1010590). Briefly, reads were aligned to yeast genome (www.yeastgenome.com) by BWA software. Single base substitutions were detected using Samtools and VarScan. Twenty individual recombinants (labeled W1_CEN to W20_CEN) were analyzed. The specific properties of each recombinant are described in Table S3 of a manuscript submitted for publication by Kozmin et al. To understand the toxic mechanisms of furfural-induced genomic instability, whole-genome sequencing of yeast strains was performed on the Illumina HiSeq 2500 sequencer using a paired-end indexing protocol. Mapping of the sequencing reads and point mutation calling were performed as described in our previous study. Briefly, reads were aligned to yeast genome (www.yeastgenome.com) by BWA software. Single base substitutions were detected using Samtools and VarScan. 21 yeast strains which were treated with 2% furfural for 2h repeated for 18 generations to accumulate genomic alterations. For each generation, after furfural treatment, cells were streaked on YPD plates and cultured at 30 degree for 2days to form single colony for the next round furfural treatment. 4 out of these 21 isolates were analyzed by Illumina high-throughput sequencing (HTS).