Journal: NAR Cancer

Article Title: Single-cell multi-gene identification of somatic mutations and gene rearrangements in cancer

doi: 10.1093/narcan/zcad034

Figure Lengend Snippet: Cancer samples used for single-cell adaptive sequencing

Article Snippet: The libraries were prepared with 900 fmol of pooled amplicon for PromethION Flow Cell FLO-PRO002 (Oxford Nanopore Technologies) using Native Barcoding Expansion and Ligation Sequencing Kit (Oxford Nanopore Technologies) as per the manufacturer’s protocol.

Techniques: Mutagenesis, Variant Assay, CRISPR, Sequencing, Transduction, Amplification

Journal: NAR Cancer

Article Title: Single-cell multi-gene identification of somatic mutations and gene rearrangements in cancer

doi: 10.1093/narcan/zcad034

Figure Lengend Snippet: An adaptive sampling method for sequencing target cDNAs from scRNA-seq. ( A ) Overview of single-cell library preparation, long- and short-read sequencing analysis, and integration of results from both modalities. Integrative Genomics Viewer (IGV) screenshot of SRSF5 targeting sites from cells with gRNAs: ( B ) SRSF5-1 and ( C ) SRSF5-2. ( D ) Boxplot showing CRISPR-induced mutation rate for all genes targeted.

Article Snippet: The libraries were prepared with 900 fmol of pooled amplicon for PromethION Flow Cell FLO-PRO002 (Oxford Nanopore Technologies) using Native Barcoding Expansion and Ligation Sequencing Kit (Oxford Nanopore Technologies) as per the manufacturer’s protocol.

Techniques: Sampling, Sequencing, CRISPR, Mutagenesis

Journal: NAR Cancer

Article Title: Single-cell multi-gene identification of somatic mutations and gene rearrangements in cancer

doi: 10.1093/narcan/zcad034

Figure Lengend Snippet: Single-cell mutations from the T1 and T2 appendiceal cancers. ( A ) Location of tumor samples for patient 8605, and variants detected from clinical diagnostic sequencing having sufficient long-read depth for analysis. ( B ) IGV screenshots for T1 alignments, covering the lengths of KRAS and GNAS genes. ( C ) UMAP clustered plot showing integration of T1 and T2 samples. ( D ) UMAP clustered plot annotated with cell types and dot plot showing expression of cell type markers. ( E ) IGV screenshot of T1 and T2 alignments showing GNAS R844S mutation position, UMAP plot highlighting location of cells with GNAS mutation and violin plot showing relative expression of mutated and wild-type GNAS epithelial cells. ( F ) IGV screenshot of T1 and T2 alignments showing KRAS G12V mutation position, UMAP plot highlighting location of cells with KRAS mutation and violin plot showing relative expression of mutated and wild-type KRAS epithelial cells.

Article Snippet: The libraries were prepared with 900 fmol of pooled amplicon for PromethION Flow Cell FLO-PRO002 (Oxford Nanopore Technologies) using Native Barcoding Expansion and Ligation Sequencing Kit (Oxford Nanopore Technologies) as per the manufacturer’s protocol.

Techniques: Diagnostic Assay, Sequencing, Expressing, Mutagenesis

Journal: NAR Cancer

Article Title: Single-cell multi-gene identification of somatic mutations and gene rearrangements in cancer

doi: 10.1093/narcan/zcad034

Figure Lengend Snippet: Single-cell mutations from the T3 and T4 appendiceal cancers. ( A ) Location of tumor samples for patient 8629, and variants detected from clinical diagnostic sequencing having sufficient long-read depth for analysis. ( B ) IGV screenshots for T3 alignments, covering the length of KRAS and GNAS genes. ( C ) UMAP clustered plot showing integration with T3 and T4 samples. ( D ) UMAP clustered plot annotated with cell types and dot plot showing expression of cell type markers. ( E ) IGV screenshot of T3 and T4 alignments showing KRAS G12V mutation position, UMAP plot highlighting location of cells with KRAS mutation and violin plot showing relative expression of mutated and wild-type KRAS epithelial cells. ( F ) IGV screenshot of T3 and T4 alignments showing GNAS R844C and R844H mutation positions, UMAP plot highlighting location of cells with GNAS R844H mutation and violin plot showing relative expression of mutated and wild-type GNAS R844H epithelial cells.

Article Snippet: The libraries were prepared with 900 fmol of pooled amplicon for PromethION Flow Cell FLO-PRO002 (Oxford Nanopore Technologies) using Native Barcoding Expansion and Ligation Sequencing Kit (Oxford Nanopore Technologies) as per the manufacturer’s protocol.

Techniques: Diagnostic Assay, Sequencing, Expressing, Mutagenesis

Journal: NAR Cancer

Article Title: Single-cell multi-gene identification of somatic mutations and gene rearrangements in cancer

doi: 10.1093/narcan/zcad034

Figure Lengend Snippet: Single-cell mutations from the T5 and T6 B-cell lymphomas. ( A ) Location of tumor samples for patient 6408 and location of biopsy taken for clinical diagnostic sequencing, plus mutations detected from targeted sequencing having sufficient long-read depth for analysis. ( B ) IGV screenshots for T5 alignments, covering the length of CREBBP and BCL2 genes. ( C ) UMAP clustered plot showing integration of T5 and T6 samples. ( D ) UMAP clustered plot annotated with cell types and dot plot showing expression of cell type markers. ( E ) IGV screenshot of T5 and T6 alignments showing BCL2 mutation positions, UMAP plot highlighting location of cells with BCL2 S116F mutation and violin plot showing relative expression of mutated and wild-type BCL2 S116F B cells, with an asterisk indicating significant difference in expression (adjusted P -value <0.05). ( F ) IGV screenshot of T5 and T6 alignments showing CREBBP Y1482H mutation position, UMAP plot highlighting location of cells with CREBBP mutation and violin plot showing relative expression of mutated and wild-type CREBBP B cells.

Article Snippet: The libraries were prepared with 900 fmol of pooled amplicon for PromethION Flow Cell FLO-PRO002 (Oxford Nanopore Technologies) using Native Barcoding Expansion and Ligation Sequencing Kit (Oxford Nanopore Technologies) as per the manufacturer’s protocol.

Techniques: Diagnostic Assay, Sequencing, Expressing, Mutagenesis

Journal: NAR Cancer

Article Title: Single-cell multi-gene identification of somatic mutations and gene rearrangements in cancer

doi: 10.1093/narcan/zcad034

Figure Lengend Snippet: ( A ) IGV screenshots of T5 and T6 lymphomas and locations of BCL2 variants called by Longshot. Coding mutations are labeled in gray and additional variants detected by Longshot are labeled in blue. ( B ) Schematic of translocation detected by cuteSV. An IGV screenshot showing primary alignments to IGH-D2 on chromosome 14 with soft-clipped sequence to the right, plus secondary alignments of the same reads to a region downstream of BCL2 3′ UTR on chromosome 18.

Article Snippet: The libraries were prepared with 900 fmol of pooled amplicon for PromethION Flow Cell FLO-PRO002 (Oxford Nanopore Technologies) using Native Barcoding Expansion and Ligation Sequencing Kit (Oxford Nanopore Technologies) as per the manufacturer’s protocol.

Techniques: Labeling, Translocation Assay, Sequencing