Journal: eLife

Article Title: Expandable and reversible copy number amplification drives rapid adaptation to antifungal drugs

doi: 10.7554/eLife.58349

Figure Lengend Snippet: ( A ) Whole genome sequence data of progenitor isolate (AMS3050) and miconazole-evolved single colonies (AMS3053, AMS3054, AMS3052, and AMS3051) plotted as in . All four colonies are monosomic from the Chr3L telomere to a CNV breakpoint at Repeat 124 (blue lines). Stair-step complex CNVs (3 to 14 copies per genome) occur on Chr3L between two distinct long inverted repeat sequences: Repeat 124 (blue lines) and Repeat 127 (orange lines) (detailed in ). All four colonies are trisomic for the Chr3 centromere ( CEN3 , black circle) and all of Chr3R. ( B ) The MIC 50 increases with the copy number of the complex CNV. Heat map of isolate growth (OD 600 at 48 hr) in twofold increasing concentrations of miconazole. The MIC 50 is denoted with a yellow line. Each heat map represents the average of three independent MIC 50 assays . ( C ) Schematic of the homologous chromosomes in AMS3051. The full length (gray) and dicentric CNV-containing (black) homologs with the positions indicated for CEN3 (circle), the long repeat sequences (blue and orange lines), and SacII cut sites (dashed lines). Four regions ( I–IV ) that support a breakage-fusion-bridge mechanism for the formation of complex CNVs (see main text for details). ( D ) Allele ratio plot of all heterozygous loci located within and flanking the complex CNV for AMS3050 and AMS3051. Allele ratio plots for all isolates are in . ( E ) SacII-digested CHEF karyotype of the progenitor and miconazole-evolved isolates (first lane of undigested AMS3050 is shown for relative size). The SacII digest isolates the region with variable copy number and CEN3 (schematic at far right). CHEF gel stained with ethidium bromide (left panel) and analyzed by Southern blot using a DIG-labeled probe to orf19.344, located within the complex CNV (middle panel), and CEN3 (right panel). Both Southern blot probes detect a novel band that increases in size as the complex CNV increases in copy number.

Article Snippet: Oxford Nanopore minion Fastq files from AMS3051 were aligned to the SC5314 reference genome (A21-s02-m09-r08) using NGMLR (-x ont, v0.2.7) ( ).

Techniques: Sequencing, Staining, Southern Blot, Labeling

Journal: eLife

Article Title: Expandable and reversible copy number amplification drives rapid adaptation to antifungal drugs

doi: 10.7554/eLife.58349

Figure Lengend Snippet: Relative genome sequence read depth plotted according to chromosome position for ( A ) the progenitor AMS3050 and ( B ) the miconazole-evolved isolate AMS3051. The complex CNV in AMS3051 is flanked by two, distinct long inverted repeat sequences: Repeat 124 (blue arrows/data points) and Repeat 127 (orange arrows/data points). The highest copy number amplification occurs between the two non-homologous inverted repeats. All features of the CNV breakpoints are detailed in ; repeat numbers refer to from .

Article Snippet: Oxford Nanopore minion Fastq files from AMS3051 were aligned to the SC5314 reference genome (A21-s02-m09-r08) using NGMLR (-x ont, v0.2.7) ( ).

Techniques: Sequencing, Amplification

Journal: eLife

Article Title: Expandable and reversible copy number amplification drives rapid adaptation to antifungal drugs

doi: 10.7554/eLife.58349

Figure Lengend Snippet: Allele ratio plot of all heterozygous loci located within and flanking the CNV breakpoints (blue and orange arrows indicate the inverted repeat sequences at the CNV breakpoints) and the CEN3 (black dot). Allele ratios shift from +0.52/- 0.48 in the progenitor AMS3050 to +0.66/- 0.34 in AMS3053, +0.77/- 0.23 in AMS3054, +0.79/- 0.21 in AMS3052, and 0.90+/- 0.10 in the 14 copy CNV region of the evolved isolate AMS3051. The specific C. albicans chromosome A/B haplotypes were reported previously .

Article Snippet: Oxford Nanopore minion Fastq files from AMS3051 were aligned to the SC5314 reference genome (A21-s02-m09-r08) using NGMLR (-x ont, v0.2.7) ( ).

Techniques:

Journal: eLife

Article Title: Expandable and reversible copy number amplification drives rapid adaptation to antifungal drugs

doi: 10.7554/eLife.58349

Figure Lengend Snippet: ( A ) Relative Illumina read depth for the miconazole-evolved isolate AMS3051 plotted by chromosome position (data as in ) indicating the presence of two inverted repeats (Repeat 124 (blue line) and Repeat 127 (orange line)) flanking a complex CNV on Chr3L. Repeat numbers refer to from . In the reference genome, Repeat 124 ( B ), consists of two inverted copies (~99% sequence identity) that are ~3 kb in length that are ~11 kb apart and Repeat 127 ( C ) consists of two inverted copies (~99% sequence identity) that are ~4 kb in length and located ~100 kb apart. Analysis of long-read sequences from AMS3051 identified structural variants relative to the reference genome at both Repeats 124 and 127. A single representative long-read (bottom purple line) aligned to the reference genome (top black line) is shown for Repeat 124 ( B ) and Repeat 127 ( C ). The long-read contains unique sequences that are separated by up to ~100 kb in the reference genome. Green-colored areas indicate alignment to the complement strand and gray-colored areas indicate alignment to the reverse complement strand of the reference genome. The transition between complement/reverse complement occurs within the repeat sequence. Schematics of both Repeat 124 and Repeat 127 indicate the formation of a fold-back inversion and non-allelic homologous recombination (NAHR, black and purple dashed lines) between repeat copies on sister chromatids, which could generate a dicentric chromosome. Alignment of the recombination product is inferred to produce the long-read that was detected (purple bar within the dicentric chromosomes). Each structural variant was supported by ~50 long-read sequences (see Materials and methods). All features of the CNV breakpoints are detailed in .

Article Snippet: Oxford Nanopore minion Fastq files from AMS3051 were aligned to the SC5314 reference genome (A21-s02-m09-r08) using NGMLR (-x ont, v0.2.7) ( ).

Techniques: Sequencing, Homologous Recombination, Variant Assay

Journal: eLife

Article Title: Expandable and reversible copy number amplification drives rapid adaptation to antifungal drugs

doi: 10.7554/eLife.58349

Figure Lengend Snippet: ( A ) Representative images of the progenitor (AMS3050) and the four miconazole-evolved isolates (AMS3053, AMS3054, AMS3052, and AMS3051) containing the Chr3L CNV grown on YPAD. Copy number of Chr3 (from ) shown to the right of the plate images. The notch in Chr3 is CEN3 . Representative images below AMS3051 of single colonies derived from either a small (blue) or large (black) colony of AMS3051 on rich medium: The small colony gave rise to AMS3094 and AMS3093 and the large colony gave rise to AMS3092. Copy number of Chr3 shown below the plate images. Whole genome sequencing data are provided in . ( B ) Colony size analysis using ImageJ (see Materials and methods), n > 113 (up to n = 300) single colonies, three biological replicates. ( C ) Heat map of OD 600 values taken at 48 hr in twofold increasing concentrations of miconazole. The MIC 50 is denoted with a yellow line. Each heat map represents the average of three independent MIC 50 assays . ( D ) CHEF of the parental strain and miconazole-evolved isolates. Whole genomic DNA was digested with SacII to isolate the region containing the CNV and CEN3 (schematic to right). CHEF gel stained with ethidium bromide (left panel) and analyzed by Southern blot with DIG-labeled probes to orf19.344 within the CNV (middle panel) and CEN3 (right panel).

Article Snippet: Oxford Nanopore minion Fastq files from AMS3051 were aligned to the SC5314 reference genome (A21-s02-m09-r08) using NGMLR (-x ont, v0.2.7) ( ).

Techniques: Derivative Assay, Sequencing, Staining, Southern Blot, Labeling

Journal: eLife

Article Title: Expandable and reversible copy number amplification drives rapid adaptation to antifungal drugs

doi: 10.7554/eLife.58349

Figure Lengend Snippet: Whole genome sequence data of miconazole-evolved isolates plotted as log2 ratio and converted to chromosome copy number (y-axis) and chromosome position (x-axis) using the Yeast Mapping Analysis Pipeline (YMAP). ( A ) The progenitor strain (AMS3050) and the four evolved isolates (AMS3053, AMS3054, AMS3052, and AMS3051) with the Chr3 CNV and Chr7 trisomy. MIC 50 at 48 hr to miconazole shown to the right. ( B ) The three single colonies (AMS3094, AMS3092, and AMS3093) derived from AMS3051. MIC 50 shown to the right.

Article Snippet: Oxford Nanopore minion Fastq files from AMS3051 were aligned to the SC5314 reference genome (A21-s02-m09-r08) using NGMLR (-x ont, v0.2.7) ( ).

Techniques: Sequencing, Derivative Assay

Journal: eLife

Article Title: Expandable and reversible copy number amplification drives rapid adaptation to antifungal drugs

doi: 10.7554/eLife.58349

Figure Lengend Snippet: Allele ratio plots of all heterozygous loci on Chr3 for ( A ) the CNV-containing progenitor isolate AMS3051, and three single colonies isolated from AMS3051 in the absence of antifungal drug ( B–D ). Location of the long inverted repeats (blue and orange arrows) and CEN3 (black dot) are shown above each plot. The progenitor to AMS3051 is homozygous on Chr3R from position 896,538 to the Chr3R telomere. ( B ) AMS3094 maintains the entire Chr3 CNV structure and allele frequency ratios similar to AMS3051. ( C ) AMS3092, which kept the complex stair-step CNV on Chr3L but became disomic for both Chr3R and the telomere proximal region of Chr3L, returned to a ~50% allele frequency ratio between the CNV and CEN3 while remaining homozygous for the telomere proximal disomic region of Chr3L. ( D ) Recombination near CEN3 resulted in LOH and loss of the CNV. The euploid strain AMS3093 became homozygous for the entire left arm of Chr3 while remaining heterozygous to the right of CEN3 .

Article Snippet: Oxford Nanopore minion Fastq files from AMS3051 were aligned to the SC5314 reference genome (A21-s02-m09-r08) using NGMLR (-x ont, v0.2.7) ( ).

Techniques: Isolation

Journal: NPJ Precision Oncology

Article Title: Benchmarking mouse contamination removing protocols in patient-derived xenografts genomic profiling

doi: 10.1038/s41698-025-00902-z

Figure Lengend Snippet: Overview of the tools benchmarked in this study

Article Snippet: Xenome , Fastq , NA , Fastq , K-mer , C + + , 2012.

Techniques: