New library construction method for single-cell genomes

Liu, Xi; Belyaev, A. N.; Spurgeon, Sandra L.; Wang, Xiaohui; Gong, Haibiao; Aboukhalil, Robert; Fekete, Richard A.

doi:10.1371/journal.pone.0181163

Cited by 22 publications

(31 citation statements)

References 34 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…Despite the promise of joint gDNA/mRNA-seq methodology, previous efforts 8,9 have suffered from limitations related to technical biases, high cost or low specificity, and have not yet seen widespread adoption. On the other hand, microwell and droplet-based ultra-low coverage genomic sequencing methods have seen major methodological improvements recently, including the implementation of direct nuclear tagmentation strategies [10][11][12][13] . These DNA-only methods focus on optimizing library preparation throughput, enabling many cells from a single sample to be analyzed with the help of specialized equipment.…”

Section: Mainmentioning

confidence: 99%

A highly scalable method for joint whole genome sequencing and gene expression profiling of single cells

Zachariadis

Andrews

et al. 2020

Preprint

View full text Add to dashboard Cite

Understanding how genetic variation alters gene expression -how genotype affects phenotype -is a central challenge in biology. To address this question in complex cell mixtures, we developed Direct Nuclear Tagmentation and RNA-sequencing (DNTR-seq), which enables whole genome and mRNA sequencing jointly in single cells. When applied to biobanked leukemia samples, DNTR-seq readily identified minor subclones within patients, as well as cell-type specific gene editing such as T-cell receptor rearrangements. mRNA-seq quality is equal to RNA-only methods, and the high yield combined with low positional bias of the genomic library preparation allows detection of sub-megabase aberrations at ultra low coverage of 0.5-3M read pairs per genome. Since each cell library is individually addressable, rare subpopulations can be re-sequenced at increased depth, allowing multi-tiered study designs where depth of sequencing is informed by previous results. In addition, the direct tagmentation protocol enables coverage-independent estimation of ploidy, which can be used to unambiguously identify cell singlets. Thus, DNTR-seq directly links each cell's state to its corresponding genome at a scale enabling routine analysis of heterogeneous tumors and other complex tissues.

show abstract

Section: Mainmentioning

confidence: 99%

A highly scalable method for joint whole genome sequencing and gene expression profiling of single cells

Zachariadis

Andrews

et al. 2020

Preprint

View full text Add to dashboard Cite

show abstract

“…The third experiment was designed to assess the performance of each method under different coverage variabilities. In particular, we simulated single cells whose coverage variabilities mimic those produced by three single-cell sequencing technologies (MALBAC, DOP-PCR and TnBC) [49] that have been used for CNA detection.…”

Section: Simulationsmentioning

confidence: 99%

“…Since Lorenz curves have been used to evaluate the variability of read counts [35,49,61], we used the Lorenz curves reported in [49] for simulating variabilities at different levels. We sampled the read counts for each bin by the distribution (Beta distribution) corresponding to their Lorenz curves using a Markov Chain Metropolis-Hastings method (Additional file 1: Figure S19 shows the Lorenz curves The Beta-splitting model For generating the underlying evolutionary trees, we followed a generalization of the Blum-François Beta-splitting model [62] which is inspired by Aldous' Beta-splitting model [63].…”

Section: Parameters For Varying the Read Count Distributionmentioning

confidence: 99%

Methods for Copy Number Aberration Detection from Single-cell DNA Sequencing Data

Fan¹,

Edrisi²,

Navin

et al. 2019

Preprint

View full text Add to dashboard Cite

Background: Accurate detection of copy number aberrations (CNA) can aid in understanding the genetic causes of diseases. Three methods (CopyNumber, Ginkgo, and HMMcopy) have been applied to single-cell DNA sequencing data for CNA detection. Results:In this paper, we benchmarked these three methods on simulated as well as biological datasets. We found that HMMcopy has the best accuracy of the three methods in terms of breakpoint detection but that Ginkgo is better in terms of detecting the actual copy numbers. In terms of computational requirements, HMMcopy consumes the least memory, but is in between the two other methods in terms of running time.Conclusion: While single-cell DNA sequencing is very promising for elucidating and understanding CNAs, even the best existing method does not exceed 80% accuracy. New methods that significantly improve upon the accuracy of these three methods are needed. Furthermore, with the large datasets being generated, the methods must be computationally efficient.

show abstract

“…4), to be followed by overall net chromosome copy losses 4,45 . Final proof for early whole genome duplication may be obtained using digital NGS based on Barcode-In-Genome technology on many individual GCNIS to determine actual chromosome copy numbers 46 . Subsequently, gain of 12p copies (which may be dynamic in subclones, details Supplementary Fig.…”

Section: Discussionmentioning

confidence: 99%

Molecular Heterogeneity and Early Metastatic Clone Selection in Testicular Germ Cell Cancer Development

Dorssers

Gillis

Stoop

et al. 2018

Preprint

View full text Add to dashboard Cite

Novelty and Impact: The nonseminoma subtype of testicular germ cell cancer is characterized by totipotency of the cancer stem cell which may differentiate into all embryonal tissue lineages.We investigated a large series of purified tumor cell samples of four nonseminoma cases using different omics methods. We demonstrate that intratumoral heterogeneity exists among the cancer stem cells and the histological components. Furthermore, metastases appeared to be derived from cancer stem cells not identified in the primary tumor.Keywords (3-5): Tumor Initiation; Genome Duplication; Copy Number Alteration; Cancer Progression, Cancer Stem Cell Abbreviations used: CG: Complete Genomics Inc.; ChC: choriocarcinoma; CNA: copy number alterations; dAP: direct alkaline phosphatase; EB: embryonal bodies; EC: embryonal carcinoma; FISH: fluorescent in situ hybridization; GCNIS: germ cell neoplasia in situ; IHC: immunohistochemistry; LAF: lesser allele frequency; LOH: loss of heterozygosity; NAP: nonmalignant adjacent parenchyma; NS: nonseminoma ; PGC: primordial germ cells; SE: seminoma; SNP: single nucleotide polymorphism; SNV: somatic DNA variants; TE: teratoma; TGCC: testicular germ cell cancer; TGCT: testicular germ cell tumor; WGS: whole genome sequencing; YST: yolk sac tumor.All rights reserved. No reuse allowed without permission.was not peer-reviewed) is the author/funder, who has granted bioRxiv a license to display the preprint in perpetuity.The copyright holder for this preprint (which . http://dx.doi.org/10.1101/385807 doi: bioRxiv preprint first posted online Aug. 8, 2018; 3 Abstract Testicular germ cell cancer (TGCC) is initiated during early life from a totipotent embryonic germ cell, and the most frequent malignant cancer in young Caucasian males. The goal of this study is to determine the intratumor heterogeneity, and to unravel tumor progression from initiation till therapy-resistant metastasis. In this study, we have investigated 42 purified samples of four cases of nonseminoma with intrinsic resistance to chemotherapy including different histological elements, metastatic specimens and the precursor cancer stem cells (germ cell neoplasia in situ, GCNIS) using whole genome-, and targeted sequencing. Sequence data were used to reconstruct the evolution of these cancers. Intratumor molecular heterogeneity was observed and did not correspond to the supposed histological evolution of the primary tumor. Metastases after systemic treatment were derived from cancer stem cells frequently not identified in the primary cancer. The GCNIS mostly lacked the molecular marks of the primary TGCC and comprised dominant clones that had failed to progress into a manifest malignancy. A BRCA-like mutational signature was found without evidence for direct involvement of BRCA1 and BRCA2

show abstract

New library construction method for single-cell genomes

Cited by 22 publications

References 34 publications

A highly scalable method for joint whole genome sequencing and gene expression profiling of single cells

A highly scalable method for joint whole genome sequencing and gene expression profiling of single cells

Methods for Copy Number Aberration Detection from Single-cell DNA Sequencing Data

Molecular Heterogeneity and Early Metastatic Clone Selection in Testicular Germ Cell Cancer Development

Contact Info

Product

Resources

About