scholarly journals Correcting palindromes in long reads after whole-genome amplification

2017 ◽  
Author(s):  
Sven Warris ◽  
Elio Schijlen ◽  
Henri van de Geest ◽  
Rahulsimham Vegesna ◽  
Thamara Hesselink ◽  
...  
Keyword(s):  
Whole Genome Amplification ◽  
De Novo ◽  
Single Cells ◽  
Whole Genome ◽  
Read Mapping ◽  
Genome Amplification ◽  
Long Reads ◽  
Oxford Nanopore ◽  
Long Read ◽  
Real World Datasets

AbstractNext-generation sequencing requires sufficient DNA to be available. If limited, whole-genome amplification is applied to generate additional amounts of DNA. Such amplification often results in many chimeric DNA fragments, in particular artificial palindromic sequences, which limit the usefulness of long reads from technologies such as PacBio and Oxford Nanopore. Here, we present Pacasus, a tool for correcting such errors in long reads. We demonstrate on two real-world datasets that it markedly improves subsequent read mapping and de novo assembly, yielding results similar to these that would be obtained with non-amplified DNA. With Pacasus long-read technologies become readily available for sequencing targets with very small amounts of DNA, such as single cells or even single chromosomes.

scholarly journals Correcting palindromes in long reads after whole-genome amplification

BMC Genomics ◽  
2018 ◽  
Vol 19 (1) ◽  
Author(s):  
Sven Warris ◽  
Elio Schijlen ◽  
Henri van de Geest ◽  
Rahulsimham Vegesna ◽  
Thamara Hesselink ◽  
...  
Keyword(s):  
Whole Genome Amplification ◽  
Whole Genome ◽  
Genome Amplification ◽  
Long Reads

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 De novo genome assembly of the olive fruit fly (Bactrocera oleae) developed through a combination of linked-reads and long-read technologies

2018 ◽  
Author(s):  
Haig Djambazian ◽  
Anthony Bayega ◽  
Konstantina T. Tsoumani ◽  
Efthimia Sagri ◽  
Maria-Eleni Gregoriou ◽  
...  
Keyword(s):  
Y Chromosome ◽  
De Novo ◽  
Fruit Fly ◽  
Bactrocera Oleae ◽  
Olive Fruit Fly ◽  
Olive Fruit ◽  
Long Reads ◽  
Oxford Nanopore ◽  
Long Read ◽  
Oxford Nanopore Technologies

AbstractLong-read sequencing has greatly contributed to the generation of high quality assemblies, albeit at a high cost. It is also not always clear how to combine sequencing platforms. We sequenced the genome of the olive fruit fly (Bactrocera oleae), the most important pest in the olive fruits agribusiness industry, using Illumina short-reads, mate-pairs, 10x Genomics linked-reads, Pacific Biosciences (PacBio), and Oxford Nanopore Technologies (ONT). The 10x linked-reads assembly gave the most contiguous assembly with an N50 of 2.16 Mb. Scaffolding the linked-reads assembly using long-reads from ONT gave a more contiguous assembly with scaffold N50 of 4.59 Mb. We also present the most extensive transcriptome datasets of the olive fly derived from different tissues and stages of development. Finally, we used the Chromosome Quotient method to identify Y-chromosome scaffolds and show that the long-reads based assembly generates very highly contiguous Y-chromosome assembly.JR is a member of the MinION Access Program (MAP) and has received free-of-charge flow cells and sequencing kits from Oxford Nanopore Technologies for other projects. JR has had no other financial support from ONT.AB has received re-imbursement for travel costs associated with attending Nanopore Community meeting 2018, a meeting organized my Oxford Nanopore Technologies.

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