scholarly journals Construction of thousands of single cell genome sequencing libraries using combinatorial indexing

2016 ◽  
Author(s):  
Sarah A. Vitak ◽  
Kristof A. Torkenczy ◽  
Jimi L. Rosenkrantz ◽  
Andrew J. Fields ◽  
Lena Christiansen ◽  
...  
Keyword(s):  
Single Cell ◽  
Genome Sequencing ◽  
Copy Number ◽  
Low Cost ◽  
Single Cells ◽  
Copy Number Variant ◽  
Tumor Evolution ◽  
Somatic Variation ◽  
Cell Genome ◽  
Single Cell Genome

AbstractSingle cell genome sequencing has proven to be a valuable tool for the detection of somatic variation, particularly in the context of tumor evolution and neuronal heterogeneity. Current technologies suffer from high per-cell library construction costs which restrict the number of cells that can be assessed, thus imposing limitations on the ability to quantitatively measure genomic heterogeneity within a tissue. Here, we present Single cell Combinatorial Indexed Sequencing (SCI-seq) as a means of simultaneously generating thousands of low-pass single cell libraries for the purpose of somatic copy number variant detection. In total, we constructed libraries for 16,698 single cells from a combination of cultured cell lines, frontal cortex tissue from Macaca mulatta, and two human adenocarcinomas. This novel technology provides the opportunity for low-cost, deep characterization of somatic copy number variation in single cells, providing a foundational knowledge across both healthy and diseased tissues.

scholarly journals Author response: Novel insights into breast cancer copy number genetic heterogeneity revealed by single-cell genome sequencing

2020 ◽  
Author(s):  
Timour Baslan ◽  
Jude Kendall ◽  
Konstantin Volyanskyy ◽  
Katherine McNamara ◽  
Hilary Cox ◽  
...  
Keyword(s):  
Breast Cancer ◽  
Single Cell ◽  
Genome Sequencing ◽  
Genetic Heterogeneity ◽  
Copy Number ◽  
Author Response ◽  
Cell Genome ◽  
Single Cell Genome

scholarly journals Resource: Scalable whole genome sequencing of 40,000 single cells identifies stochastic aneuploidies, genome replication states and clonal repertoires

2018 ◽  
Author(s):  
Emma Laks ◽  
Hans Zahn ◽  
Daniel Lai ◽  
Andrew McPherson ◽  
Adi Steif ◽  
...  
Keyword(s):  
Single Cell ◽  
Genome Sequencing ◽  
Large Scale ◽  
Single Cells ◽  
Genome Structure ◽  
Cancer Tissue ◽  
Genome Replication ◽  
Wide Range ◽  
Cell Genome ◽  
Single Cell Genome

SummaryEssential features of cancer tissue cellular heterogeneity such as negatively selected genome topologies, sub-clonal mutation patterns and genome replication states can only effectively be studied by sequencing single-cell genomes at scale and high fidelity. Using an amplification-free single-cell genome sequencing approach implemented on commodity hardware (DLP+) coupled with a cloud-based computational platform, we define a resource of 40,000 single-cell genomes characterized by their genome states, across a wide range of tissue types and conditions. We show that shallow sequencing across thousands of genomes permits reconstruction of clonal genomes to single nucleotide resolution through aggregation analysis of cells sharing higher order genome structure. From large-scale population analysis over thousands of cells, we identify rare cells exhibiting mitotic mis-segregation of whole chromosomes. We observe that tissue derived scWGS libraries exhibit lower rates of whole chromosome anueploidy than cell lines, and loss of p53 results in a shift in event type, but not overall prevalence in breast epithelium. Finally, we demonstrate that the replication states of genomes can be identified, allowing the number and proportion of replicating cells, as well as the chromosomal pattern of replication to be unambiguously identified in single-cell genome sequencing experiments. The combined annotated resource and approach provide a re-implementable large scale platform for studying lineages and tissue heterogeneity.

scholarly journals Massively parallel single-cell genome sequencing enables high-resolution analysis of soil and marine microbiome

2020 ◽  
Author(s):  
Yohei Nishikawa ◽  
Masato Kogawa ◽  
Masahito Hosokawa ◽  
Katsuhiko Mineta ◽  
Kai Takahashi ◽  
...  
Keyword(s):  
Single Cell ◽  
Genome Sequencing ◽  
Single Cells ◽  
Gene Clusters ◽  
Sequencing Platform ◽  
High Resolution Analysis ◽  
Resolution Analysis ◽  
Single Cell Isolation ◽  
Cell Genome ◽  
Single Cell Genome

AbstractTo improve our understanding of the environmental microbiome, we developed a single-cell genome sequencing platform, named SAG-gel, which utilizes gel beads for single-cell isolation, cell lysis, and whole genome amplification (WGA) for sequencing. SAG-gel enables serial, parallel and independent reactions of > 100,000 single cells in a single tube, delivering high-quality genome recovery with storable randomized single-cell genome libraries. From soil and marine environmental sources, we acquired 734 partial genomes that are recapitulated in 231 species, 35% of which were assigned as high-to-medium qualities. We found that each genome to be almost unique and 98.7% of them were newly identified, implying the complex genetic diversities across 44 phyla. The various metabolic capabilities including virulence factors and biosynthetic gene clusters were found across the lineages at single-cell resolution. This technology will accelerate the accumulation of reference genomes of uncharacterized environmental microbes and provide us new insights for their roles.

Characterization of Neonatal Lung Cells Through Single Cell Genome Sequencing

2019 ◽  
Author(s):  
S. Bhattacharya ◽  
J. Lillis ◽  
C. Baker ◽  
M. Guo ◽  
J.R. Myers ◽  
...  
Keyword(s):  
Single Cell ◽  
Genome Sequencing ◽  
Lung Cells ◽  
Neonatal Lung ◽  
Cell Genome ◽  
Single Cell Genome

scholarly journals MBRS-59. SINGLE-CELL WHOLE-GENOME SEQUENCING DISSECTS INTRA-TUMOURAL GENOMIC HETEROGENEITY AND CLONAL EVOLUTION IN CHILDHOOD MEDULLOBLASTOMA

2020 ◽  
Vol 22 (Supplement_3) ◽  
pp. iii408-iii408
Author(s):  
Marina Danilenko ◽  
Masood Zaka ◽  
Claire Keeling ◽  
Stephen Crosier ◽  
Rafiqul Hussain ◽  
...  
Keyword(s):  
Whole Genome Sequencing ◽  
Single Cell ◽  
Genome Sequencing ◽  
Copy Number ◽  
Single Cell Analysis ◽  
Mutational Analysis ◽  
Single Cells ◽  
Clonal Evolution ◽  
Whole Genome ◽  
Clinically Significant

Abstract Medulloblastomas harbor clinically-significant intra-tumoral heterogeneity for key biomarkers (e.g. MYC/MYCN, β-catenin). Recent studies have characterized transcriptional heterogeneity at the single-cell level, however the underlying genomic copy number and mutational architecture remains to be resolved. We therefore sought to establish the intra-tumoural genomic heterogeneity of medulloblastoma at single-cell resolution. Copy number patterns were dissected by whole-genome sequencing in 1024 single cells isolated from multiple distinct tumour regions within 16 snap-frozen medulloblastomas, representing the major molecular subgroups (WNT, SHH, Group3, Group4) and genotypes (i.e. MYC amplification, TP53 mutation). Common copy number driver and subclonal events were identified, providing clear evidence of copy number evolution in medulloblastoma development. Moreover, subclonal whole-arm and focal copy number alterations covering important genomic loci (e.g. on chr10 of SHH patients) were detected in single tumour cells, yet undetectable at the bulk-tumor level. Spatial copy number heterogeneity was also common, with differences between clonal and subclonal events detected in distinct regions of individual tumours. Mutational analysis of the cells allowed dissection of spatial and clonal heterogeneity patterns for key medulloblastoma mutations (e.g. CTNNB1, TP53, SMARCA4, PTCH1) within our cohort. Integrated copy number and mutational analysis is underway to establish their inter-relationships and relative contributions to clonal evolution during tumourigenesis. In summary, single-cell analysis has enabled the resolution of common mutational and copy number drivers, alongside sub-clonal events and distinct patterns of clonal and spatial evolution, in medulloblastoma development. We anticipate these findings will provide a critical foundation for future improved biomarker selection, and the development of targeted therapies.

scholarly journals Single-Cell, Genome-wide Sequencing Identifies Clonal Somatic Copy-Number Variation in the Human Brain

Cell Reports ◽  
2014 ◽  
Vol 8 (5) ◽  
pp. 1280-1289 ◽  
Author(s):  
Xuyu Cai ◽  
Gilad D. Evrony ◽  
Hillel S. Lehmann ◽  
Princess C. Elhosary ◽  
Bhaven K. Mehta ◽  
...  
Keyword(s):  
Copy Number Variation ◽  
Human Brain ◽  
Single Cell ◽  
Copy Number ◽  
Genome Wide ◽  
Number Variation ◽  
Cell Genome ◽  
Single Cell Genome

scholarly journals Reconstructing mutational lineages in breast cancer by multi-patient-targeted single cell DNA sequencing

2021 ◽  
Author(s):  
Nicholas Navin ◽  
Jake Leighton ◽  
Min Hu ◽  
Emi Sei ◽  
Funda Meric-Bernstam
Keyword(s):  
Breast Cancer ◽  
Dna Sequencing ◽  
Single Cell ◽  
Copy Number ◽  
Somatic Mutations ◽  
Single Cells ◽  
Tumor Evolution ◽  
Sequencing Method ◽  
Genomic Regions ◽  
Microfluidics Platform

Single cell DNA sequencing (scDNA-seq) methods are powerful tools for profiling mutations in cancer cells, however most genomic regions characterized in single cells are non-informative. To overcome this issue, we developed a Multi-Patient-Targeted (MPT) scDNA-seq sequencing method. MPT involves first performing bulk exome sequencing across a cohort of cancer patients to identify somatic mutations, which are then pooled together to develop a single custom targeted panel for high-throughput scDNA-seq using a microfluidics platform. We applied MPT to profile 330 mutations across 23,500 cells from 5 TNBC patients, which showed that 3 tumors were monoclonal and 2 tumors were polyclonal. From this data, we reconstructed mutational lineages and identified early mutational and copy number events, including early TP53 mutations that occurred in all five patients. Collectively, our data suggests that MPT can overcome technical obstacles for studying tumor evolution using scDNA-seq by profiling information-rich mutation sites.

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