Deterministic Whole-Genome Amplification of Single Cells

2015 ◽  
pp. 69-86 ◽  
Author(s):  
Zbigniew Tadeusz Czyż ◽  
Christoph A. Klein
Keyword(s):  
Whole Genome Amplification ◽  
Single Cells ◽  
Whole Genome ◽  
Genome Amplification

scholarly journals Combined Molecular Genetic and Cytogenetic Analysis from Single Cells after Isothermal Whole-Genome Amplification

2011 ◽  
Vol 57 (7) ◽  
pp. 1032-1041 ◽  
Author(s):  
Thomas Kroneis ◽  
Jochen B Geigl ◽  
Amin El-Heliebi ◽  
Martina Auer ◽  
Peter Ulz ◽  
...  
Keyword(s):  
Single Cell ◽  
Tumor Cells ◽  
Whole Genome Amplification ◽  
Single Cells ◽  
Molecular Genetic ◽  
Target Cells ◽  
Whole Genome ◽  
Single Cell Level ◽  
Cell Level ◽  
Genome Amplification

BACKGROUND Analysis of chromosomal aberrations or single-gene disorders from rare fetal cells circulating in the blood of pregnant women requires verification of the cells' genomic identity. We have developed a method enabling multiple analyses at the single-cell level that combines verification of the genomic identity of microchimeric cells with molecular genetic and cytogenetic diagnosis. METHODS We used a model system of peripheral blood mononuclear cells spiked with a colon adenocarcinoma cell line and immunofluorescence staining for cytokeratin in combination with DNA staining with the nuclear dye TO-PRO-3 in a preliminary study to define candidate microchimeric (tumor) cells in Cytospin preparations. After laser microdissection, we performed low-volume on-chip isothermal whole-genome amplification (iWGA) of single and pooled cells. RESULTS DNA fingerprint analysis of iWGA aliquots permitted successful identification of all analyzed candidate microchimeric cell preparations (6 samples of pooled cells, 7 samples of single cells). Sequencing of 3 single-nucleotide polymorphisms was successful at the single-cell level for 20 of 32 allelic loci. Metaphase comparative genomic hybridization (mCGH) with iWGA products of single cells showed the gains and losses known to be present in the genomic DNA of the target cells. CONCLUSIONS This method may be instrumental in cell-based noninvasive prenatal diagnosis. Furthermore, the possibility to perform mCGH with amplified DNA from single cells offers a perspective for the analysis of nonmicrochimeric rare cells exhibiting genomic alterations, such as circulating tumor cells.

scholarly journals TruePrime is a novel method for whole-genome amplification from single cells based on TthPrimPol

2016 ◽  
Vol 7 (1) ◽  
Author(s):  
Ángel J. Picher ◽  
Bettina Budeus ◽  
Oliver Wafzig ◽  
Carola Krüger ◽  
Sara García-Gómez ◽  
...  
Keyword(s):  
Whole Genome Amplification ◽  
Single Cells ◽  
Whole Genome ◽  
Genome Amplification ◽  
Novel Method

scholarly journals Identification of small gains and losses in single cells after whole genome amplification on tiling oligo arrays

2009 ◽  
Vol 37 (15) ◽  
pp. e105-e105 ◽  
Author(s):  
Jochen B. Geigl ◽  
Anna C. Obenauf ◽  
Julie Waldispuehl-Geigl ◽  
Eva M. Hoffmann ◽  
Martina Auer ◽  
...  
Keyword(s):  
Whole Genome Amplification ◽  
Single Cells ◽  
Whole Genome ◽  
Genome Amplification ◽  
Gains And Losses

scholarly journals A Simple Method for Encapsulating Single Cells in Alginate Microspheres Allows for Direct PCR and Whole Genome Amplification

PLoS ONE ◽  
2015 ◽  
Vol 10 (2) ◽  
pp. e0117738 ◽  
Author(s):  
Saharnaz Bigdeli ◽  
Roger O. Dettloff ◽  
Curtis W. Frank ◽  
Ronald W. Davis ◽  
Laurel D. Crosby
Keyword(s):  
Whole Genome Amplification ◽  
Single Cells ◽  
Whole Genome ◽  
Direct Pcr ◽  
Simple Method ◽  
Genome Amplification

scholarly journals Comparison of seven single cell Whole Genome Amplification commercial kits using targeted sequencing

2017 ◽  
Author(s):  
Tamir Biezuner ◽  
Ofir Raz ◽  
Shiran Amir ◽  
Lilach Milo ◽  
Rivka Adar ◽  
...  
Keyword(s):  
Single Cell ◽  
Whole Genome Amplification ◽  
Single Cells ◽  
Targeted Sequencing ◽  
Whole Genome ◽  
Genome Coverage ◽  
Genome Amplification ◽  
Cell Experiment ◽  
Commercial Kits ◽  
Genomic Regions

AbstractAdvances in biochemical technologies have led to a boost in the field of single cell genomics. Observation of the genome at a single cell resolution is currently achieved by pre-amplification using whole genome amplification (WGA) techniques that differ by their biochemical aspects and as a result by biased amplification of the original molecule. Several comparisons between commercially available single cell dedicated WGA kits (scWGA) were performed, however, these comparisons are costly, were only performed on selected scWGA kit and more notably, are limited by the number of analyzed cells, making them limited for reproducibility analysis. We benchmarked an economical assay to compare all commercially available scWGA kits that is based on targeted sequencing of thousands of genomic regions, including highly mutable genomic regions (microsatellites), from a large cohort of human single cells (125 cells in total). Using this approach, we could analyze the genome coverage, the reproducibility of genome coverage and the error rate of each kit. Our experimental design provides an affordable and reliable comparative assay that simulates a real single cell experiment. Results demonstrate the needfor a dedicated kit selection depending on the desired single cell assay.

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