Journal: bioRxiv

Article Title: Large CRISPR-Cas-induced deletions in the oxamniquine resistance locus of the human parasite Schistosoma mansoni

doi: 10.1101/2020.05.03.074831

Figure Lengend Snippet: (A) Gene model of SULT-OR ( Smp_089320 ), indicating the position of the two exons, one intron, and UTRs, spanning 4837 bp on the reverse strand of chromosome 6. Boxes filled in dark red represent the protein coding sequence. Schistosoma mansoni (PRJEA36577). Assembly: Smansoni_v7 (GCA_000237925.3). Region: Scaffold SM_V7_6:3,183,084-3,188,924. Adapted from WormBase ParaSite 14 . (B) Nucleotide sequence of the start of exon 1 indicating location and sequence of gRNA target site, predicted double-stranded break (DSB), protospacer adjacent motif (PAM), and the PvuII restriction site. (C) Reference PCR amplicon, showing the positions of the gRNA, PAM, DSB, forward and reverse PCR primers, and forward and reverse sequence reads, as well as a SNP site found in many sequences reads. The diagrams are drawn to scale.

Article Snippet: Amplicon libraries from the samples summarised in Supplementary Table S1 were sequenced on a MiSeq Illumina sequencing platform spiked with 20-30% PhiX to generate diversity.

Techniques: Sequencing, Amplification

Journal: bioRxiv

Article Title: Large CRISPR-Cas-induced deletions in the oxamniquine resistance locus of the human parasite Schistosoma mansoni

doi: 10.1101/2020.05.03.074831

Figure Lengend Snippet: (A) Frequency of deletions in NGS sequencing data, identified with the assistance of CRISPResso in three biological replicates from adults, two from sporocysts, and three from eggs. (B) CRISPR-induced deletions in adult worms and sporocysts. The positions of deletions found by CRISPResso in the reference amplicon are indicated, in three biological replicates of CRISPR-Cas9-treated adult samples (blue lines: experiment 1, tag 5; experiment 7, tag 50; experiment 11, tag 64) and matched adult control samples (red lines: experiment 1, tag 9; experiment 7, tag 82; experiment 11, tag 21), and in two biological replicates of CRISPR-Cas9-treated sporocysts (green lines: experiment 2, tag 6; experiment 11, tag 19) and matched sporocyst controls (orange lines: experiment 2, tag 15; experiment 11, tag 43). The black arrow shows the predicted Cas9 cut site. (C) Multi-sequence alignment of SULT-OR alleles with deletions found in CRISPR-Cas9-treated adult worms that are supported by >=50 reads and span the DSB site indicated with a red line, based on one of the treated adult replicates (experiment 1, tag 5). The common 34-bp deletion is highlighted in pale pink.

Article Snippet: Amplicon libraries from the samples summarised in Supplementary Table S1 were sequenced on a MiSeq Illumina sequencing platform spiked with 20-30% PhiX to generate diversity.

Techniques: Sequencing, CRISPR, Amplification, Control

Journal: bioRxiv

Article Title: Large CRISPR-Cas-induced deletions in the oxamniquine resistance locus of the human parasite Schistosoma mansoni

doi: 10.1101/2020.05.03.074831

Figure Lengend Snippet: Deletion alleles seen in the SULT-OR gene in amplicon sequencing reads from treated (A) and control (B) adults (left), sporocysts (centre), and eggs (right), showing alleles that contain a single internal deletion and no internal insertions with respect to the reference amplicon. The y-axis shows deletion alleles sorted by the number of reads supporting them, with the alleles supported by the most reads at the bottom. Alleles supported by >500 reads in red, alleles supported by 101-500 reads in dark orange, alleles supported by 11-100 reads in pale orange, and alleles supported by 1-10 reads in pale green. The x-axis shows the position of the deletion along the reference amplicon, with a blue vertical line at the predicted Cas9 cut site.

Article Snippet: Amplicon libraries from the samples summarised in Supplementary Table S1 were sequenced on a MiSeq Illumina sequencing platform spiked with 20-30% PhiX to generate diversity.

Techniques: Amplification, Sequencing, Control