Supplemental Material for Möller et al., 2018
Table S1: List of primers used for screening for accessory chromosomes in Z. triticiand Z. ardabiliae.
Table S2: List of chromosome-loss strains identified in the in vitroand in plantaexperiments.
Table S3: Chromosome variation of sequenced Zt09-derived strains grown at 28°C.
Table S4: Location and annotation of SNPs and INDELs found in the sequenced chromosome-loss strains.
Table S5: Sequence annotation of chromosomal breakpoints detected in the sequenced chromosome-loss strains derived from the temperature stress experiment.
Figure S1: Southern blotting of pulsed-field gels using chromosome-specific probes confirms chromosome fusions. To prove and visualize chromosome fusions in the strains derived from the temperature stress experiment, we conducted PFGE followed by Southern blots with probes specific to the chromosomes, which based on genome data and karyotype analyses, were found to have fused. Panel (A) shows a color-inverted image of the pulsed-field gel (top) and Southern blot (bottom) using a probe specific for chromosome 17. The genomes of all tested strains have chromosome 17, and the genomes of strains Zt09 28-2 and Zt09 28-4 contain a duplicated chromosome 17 (Figure 2C). While in Zt09 28-2 the duplication resulted in two separate copies of the chromosome, the two chromosomes 17 in Zt09 28-4 fused and formed a new ~ 1.2 Mb chromosome. (B) For the second Southern blot, we used probes for chromosomes 12 and 21. Chromosome 21 is absent in the strains Zt09 28-1 and Zt09 28-3 (PFGE, top and Southern analyses, bottom). Genome sequencing shows that chromosome 21 is present in Zt09 28-2 (Figure 2C), however the respective band on the pulsed-field gel is missing. Similarly, chromosomes 12 and 13 are not present in this strain (PFGE, top), while the chromosome sequences are present in the whole genome sequence data. Apparent loss of chromosome 13 can be explained by the fusion of chromosomes 3 and 13, as indicated by the sequence analysis (Figure 3). The probes for chromosomes 12 and 21 both hybridize to one band in the size of ~1.8 Mb (Southern, bottom) matching the size expected if a chromosome fusion occurred and explaining the absence of their respective ‘original’ chromosomal bands.
Figure S2: Growth assay of Zt09 and chromosome-loss strains Zt09∆14 and Zt09∆21. Three independent growth assays were conducted with the progenitor strain Zt09 (A) and two chromosome-loss strains Zt09∆14 (B) and Zt09∆21 (C), representing the loss of the largest and smallest accessory chromosome. Plotted are the fitted growth curves of all replicates and experiments generated with the R package growthcurver. The growth curves are based on OD600measurements. Panel (D) displays a boxplot of all r values of the different strains. There are no significant differences between the growth curves of the three tested strains (Wilcoxon rank-sum test Zt09 – Zt09∆14: p-value = 0.1443, Zt09 - Zt09∆21: p-value = 0.3762).