Somatic variant calling from single-cell DNA sequencing data

Methods and applications for single-cell and spatial multi-omics

“…While the supernatant cytosol is isolated for Smart-seq2-like scRNA-seq, the nuclear DNA is subjected to direct tagmentation . The latter enables direct PCR-based library preparation for scDNA-seq, thereby circumventing the classic approach of WGA before scDNA-seq library preparation and in part the resulting artefacts associated with it 14 , 15 . However, these nuclear–cytosolic partitioning methods are less amenable to comprehensive characterization of mitochondrial DNA and nuclear RNA, and they are confined to the use of intact cells as input.…”

“…This principle has been shown for the detection of single nucleotide variants contained within expressed genes, forms of structural variation resulting in the expression of fusion genes, and copy number variants resulting in gene expression dosage effects. The genomic and mitochondrial DNA variants detected by genome-plus-transcriptome sequencing approaches, considering potential imperfections 15 , can furthermore be leveraged for the construction of a genetic lineage tree of the cells, which can be annotated with cell type and functional phenotypic states of cells using the RNA sequencing data of the same cells. As an alternative to analysing naturally occurring somatic mutations, high-throughput methods relying on CRISPR-scarring are available in model systems 32 , which through recent improvements may enable accurate lineage recording as well as the capturing of ancestral transcriptional states 33 , 34 .…”

Grave-to-cradle: human embryonic lineage tracing from the postmortem body

“…Several single-cell WGA methods have been developed thus far 76 , and various computational approaches have been applied to identify somatic variants 82 . Among them, the recently developed isothermal WGA method, primary template-directed amplification (PTA), is attracting attention 83 .…”

Unraveling Heterogeneity in the Aging Hematopoietic Stem Cell Compartment: An Insight From Single-cell Approaches

“…99 Many scDNA-Seq techniques are error prone due to technical biases arising from uneven sequence coverage, allelic dropout, and the introduction of errors during amplification steps in sequence library construction. 100 Despite the error-prone nature of scDNA-Seq data, genotyping studies targeting the somatic mutations of single cells in human clonal hematopoiesis and acute myeloid leukemia have revealed the dominance of a small number of clones that frequently contain co-occurring mutations in epigenetic regulators, thereby promoting clonal expansion. For example, the co-occurence of mutations in Nucleophosmin 1 and an internal tandem duplication mutation in the Flt3 gene or the combination of mutations in DNA methyltransferase 3 alpha (DNMT3A) and isocitrate dehydrogenase (NADP[+]) 2 were found to be associated with clonal dominance.…”

“…Single‐cell DNA sequencing (scDNA‐Seq) encompasses a number of techniques that have been used to detect various genomic alterations such as single nucleotide polymorphisms, insertions, and deletions 99 . Many scDNA‐Seq techniques are error prone due to technical biases arising from uneven sequence coverage, allelic dropout, and the introduction of errors during amplification steps in sequence library construction 100 . Despite the error‐prone nature of scDNA‐Seq data, genotyping studies targeting the somatic mutations of single cells in human clonal hematopoiesis and acute myeloid leukemia have revealed the dominance of a small number of clones that frequently contain co‐occurring mutations in epigenetic regulators, thereby promoting clonal expansion.…”