Towards the analysis of the genomes of single cells: Further characterisation of the multiple displacement amplification

Gene ◽  
2006 ◽  
Vol 372 ◽  
pp. 1-7 ◽  
Author(s):  
Simona Panelli ◽  
Giuseppe Damiani ◽  
Luca Espen ◽  
Gioacchino Micheli ◽  
Vittorio Sgaramella
Keyword(s):  
Multiple Displacement Amplification ◽  
Single Cells

scholarly journals Development of Microbial Genome-Probing Microarrays using Digital Multiple Displacement Amplification of Uncultivated Microbial Single Cells

2008 ◽  
Vol 42 (22) ◽  
pp. 8614-8614
Author(s):  
Ho-Won Chang ◽  
Youboong Sung ◽  
Kyoung-Ho Kim ◽  
Young-Do Nam ◽  
Seong Woon Roh ◽  
...  
Keyword(s):  
Multiple Displacement Amplification ◽  
Single Cells ◽  
Microbial Genome

scholarly journals SCELLECTOR: ranking amplification bias in single cells using shallow sequencing

2020 ◽  
Vol 21 (1) ◽  
Author(s):  
Vivekananda Sarangi ◽  
Alexandre Jourdon ◽  
Taejeong Bae ◽  
Arijit Panda ◽  
Flora Vaccarino ◽  
...  
Keyword(s):  
Single Cell ◽  
Multiple Displacement Amplification ◽  
Single Cells ◽  
Sequencing Data ◽  
High Coverage ◽  
Amplification Bias ◽  
Single Cell Profiling ◽  
High Concordance ◽  
Human Neuronal Cells

Abstract Background The study of mosaic mutation is important since it has been linked to cancer and various disorders. Single cell sequencing has become a powerful tool to study the genome of individual cells for the detection of mosaic mutations. The amount of DNA in a single cell needs to be amplified before sequencing and multiple displacement amplification (MDA) is widely used owing to its low error rate and long fragment length of amplified DNA. However, the phi29 polymerase used in MDA is sensitive to template fragmentation and presence of sites with DNA damage that can lead to biases such as allelic imbalance, uneven coverage and over representation of C to T mutations. It is therefore important to select cells with uniform amplification to decrease false positives and increase sensitivity for mosaic mutation detection. Results We propose a method, Scellector (single cell selector), which uses haplotype information to detect amplification quality in shallow coverage sequencing data. We tested Scellector on single human neuronal cells, obtained in vitro and amplified by MDA. Qualities were estimated from shallow sequencing with coverage as low as 0.3× per cell and then confirmed using 30× deep coverage sequencing. The high concordance between shallow and high coverage data validated the method. Conclusion Scellector can potentially be used to rank amplifications obtained from single cell platforms relying on a MDA-like amplification step, such as Chromium Single Cell profiling solution.

O▪28 Validating multiple displacement amplification for analysis of single cells

2005 ◽  
Vol 10 ◽  
pp. 10
Author(s):  
AR Thornhill ◽  
KV Schowalter ◽  
JR Fredrickson ◽  
DL Walker ◽  
SN Thibodeau
Keyword(s):  
Multiple Displacement Amplification ◽  
Single Cells

scholarly journals Genome sequencing and de novo assembly of the giant unicellular alga Acetabularia acetabulum using droplet MDA

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Ina J. Andresen ◽  
Russell J. S. Orr ◽  
Anders K. Krabberød ◽  
Kamran Shalchian-Tabrizi ◽  
Jon Bråte
Keyword(s):  
Genome Sequencing ◽  
Cell Biology ◽  
De Novo ◽  
Multiple Displacement Amplification ◽  
Single Cells ◽  
Model System ◽  
Genomic Information ◽  
Unicellular Alga ◽  
Acetabularia Acetabulum ◽  
Genomic Material

AbstractThe macroscopic single-celled green alga Acetabularia acetabulum has been a model system in cell biology for more than a century. However, no genomic information is available from this species. Since the alga has a long life cycle, is difficult to grow in dense cultures, and has an estimated diploid genome size of almost 2 Gb, obtaining sufficient genomic material for genome sequencing is challenging. Here, we have attempted to overcome these challenges by amplifying genomic DNA using multiple displacement amplification (MDA) combined with microfluidics technology to distribute the amplification reactions across thousands of microscopic droplets. By amplifying and sequencing DNA from five single cells we were able to recover an estimated ~ 7–11% of the total genome, providing the first draft of the A. acetabulum genome. We highlight challenges associated with genome recovery and assembly of MDA data due to biases arising during genome amplification, and hope that our study can serve as a reference for future attempts on sequencing the genome from non-model eukaryotes.

Development of Microbial Genome-Probing Microarrays Using Digital Multiple Displacement Amplification of Uncultivated Microbial Single Cells

2008 ◽  
Vol 42 (16) ◽  
pp. 6058-6064 ◽  
Author(s):  
Ho-Won Chang ◽  
Youlboong Sung ◽  
Kyoung-Ho Kim ◽  
Young-Do Nam ◽  
Seong Woon Roh ◽  
...  
Keyword(s):  
Multiple Displacement Amplification ◽  
Single Cells ◽  
Microbial Genome

scholarly journals Nanoliter Reactors Improve Multiple Displacement Amplification of Genomes from Single Cells

PLoS Genetics ◽  
2007 ◽  
Vol 3 (9) ◽  
pp. e155 ◽  
Author(s):  
Yann Marcy ◽  
Thomas Ishoey ◽  
Roger S Lasken ◽  
Timothy B Stockwell ◽  
Brian P Walenz ◽  
...  
Keyword(s):  
Multiple Displacement Amplification ◽  
Single Cells

scholarly journals Sequencing Metrics of Human Genomes Extracted from Single Cancer Cells Individually Isolated in a Valveless Microfluidic Device

2018 ◽  
Author(s):  
Rodolphe Marie ◽  
Marie Pødenphant ◽  
Kamila Koprowska ◽  
Loic Bærlocher ◽  
Roland C.M. Vulders ◽  
...  
Keyword(s):  
Microfluidic Device ◽  
Dna Sequences ◽  
Multiple Displacement Amplification ◽  
Single Cells ◽  
Direct Determination ◽  
High Quality ◽  
Human Genomes ◽  
Colorectal Cell ◽  
Single Cancer

AbstractSequencing the genomes of individual cells enables the direct determination of genetic heterogeneity amongst cells within a population. We have developed an injection-moulded valveless microfluidic device in which single cells from colorectal cell (LS174T, LS180 and RKO) lines and fresh colorectal cancers are individually trapped, their genomes extracted and prepared for sequencing, using multiple displacement amplification (MDA). Ninety nine percent of the DNA sequences obtained mapped to a reference human genome, indicating that there was effectively no contamination of these samples from non-human sources. In addition, most of the reads are correctly paired, with a low percentage of singletons (0.17 ± 0.06 %) and we obtain genome coverages approaching 90%. To achieve this high quality, our device design and process shows that amplification can be conducted in microliter volumes as long as extraction is in sub-nanoliter volumes. Our data also demonstrates that high quality single cell sequencing can be achieved using a relatively simple, inexpensive and scalable device.

scholarly journals Evaluation of genome coverage and fidelity of multiple displacement amplification from single cells by SNP array

2009 ◽  
Vol 15 (11) ◽  
pp. 739-747 ◽  
Author(s):  
J. Ling ◽  
G. Zhuang ◽  
B. Tazon-Vega ◽  
C. Zhang ◽  
B. Cao ◽  
...  
Keyword(s):  
Multiple Displacement Amplification ◽  
Single Cells ◽  
Snp Array ◽  
Genome Coverage

scholarly journals Robust high-performance nanoliter-volume single-cell multiple displacement amplification on planar substrates

2016 ◽  
Vol 113 (30) ◽  
pp. 8484-8489 ◽  
Author(s):  
Kaston Leung ◽  
Anders Klaus ◽  
Bill K. Lin ◽  
Emma Laks ◽  
Justina Biele ◽  
...  
Keyword(s):  
Ovarian Cancer ◽  
Single Cell ◽  
High Throughput ◽  
Multiple Displacement Amplification ◽  
Single Cells ◽  
Biological Significance ◽  
Phase Cell ◽  
Whole Genome ◽  
Large Numbers ◽  
Diploid Cells

The genomes of large numbers of single cells must be sequenced to further understanding of the biological significance of genomic heterogeneity in complex systems. Whole genome amplification (WGA) of single cells is generally the first step in such studies, but is prone to nonuniformity that can compromise genomic measurement accuracy. Despite recent advances, robust performance in high-throughput single-cell WGA remains elusive. Here, we introduce droplet multiple displacement amplification (MDA), a method that uses commercially available liquid dispensing to perform high-throughput single-cell MDA in nanoliter volumes. The performance of droplet MDA is characterized using a large dataset of 129 normal diploid cells, and is shown to exceed previously reported single-cell WGA methods in amplification uniformity, genome coverage, and/or robustness. We achieve up to 80% coverage of a single-cell genome at 5× sequencing depth, and demonstrate excellent single-nucleotide variant (SNV) detection using targeted sequencing of droplet MDA product to achieve a median allelic dropout of 15%, and using whole genome sequencing to achieve false and true positive rates of 9.66 × 10−6 and 68.8%, respectively, in a G1-phase cell. We further show that droplet MDA allows for the detection of copy number variants (CNVs) as small as 30 kb in single cells of an ovarian cancer cell line and as small as 9 Mb in two high-grade serous ovarian cancer samples using only 0.02× depth. Droplet MDA provides an accessible and scalable method for performing robust and accurate CNV and SNV measurements on large numbers of single cells.

Whole-genome multiple displacement amplification from single cells

2006 ◽  
Vol 1 (4) ◽  
pp. 1965-1970 ◽  
Author(s):  
Claudia Spits ◽  
Cédric Le Caignec ◽  
Martine De Rycke ◽  
Lindsey Van Haute ◽  
André Van Steirteghem ◽  
...  
Keyword(s):  
Multiple Displacement Amplification ◽  
Single Cells ◽  
Whole Genome
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