Repli-seq: genome-wide analysis of replication timing by next-generation sequencing

ABSTRACTCycling cells duplicate their DNA content during S phase, following a defined program called replication timing (RT). Early and late replicating regions differ in terms of mutation rates, transcriptional activity, chromatin marks and sub-nuclear position. Moreover, RT is regulated during development and is altered in disease. Exploring mechanisms linking RT to other cellular processes in normal and diseased cells will be facilitated by rapid and robust methods with which to measure RT genome wide. Here, we describe a rapid, robust and relatively inexpensive protocol to analyze genome-wide RT by next-generation sequencing (NGS). This protocol yields highly reproducible results across laboratories and platforms. We also provide computational pipelines for analysis, parsing phased genomes using single nucleotide polymorphisms (SNP) for analyzing RT allelic asynchrony, and for direct comparison to Repli-chip data obtained by analyzing nascent DNA by microarrays.

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Quantification of fetal DNA in the plasma of pregnant women using next generation sequencing of frequent single nucleotide polymorphisms

Bulletin of Russian State Medical University ◽

10.24075/brsmu.2018.031 ◽

2018 ◽

pp. 29-33

Author(s):

J. Shubina ◽

◽

T Jankevic ◽

A. Yu. Goltsov ◽

I. S. Mukosey ◽

...

Keyword(s):

Next Generation Sequencing ◽

Single Nucleotide Polymorphisms ◽

Pregnant Women ◽

Nucleotide Polymorphisms ◽

Next Generation ◽

Single Nucleotide ◽

Fetal Dna ◽

Generation Sequencing

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Mapping DNA Topoisomerase Binding and Cleavage Genome Wide Using Next-Generation Sequencing Techniques

Genes ◽

10.3390/genes11010092 ◽

2020 ◽

Vol 11 (1) ◽

pp. 92 ◽

Cited By ~ 1

Author(s):

Shannon J. McKie ◽

Anthony Maxwell ◽

Keir C. Neuman

Keyword(s):

Next Generation Sequencing ◽

Dna Cleavage ◽

Next Generation ◽

Dna Breaks ◽

Genome Wide ◽

Extension Activity ◽

Nucleotide Resolution ◽

Next Generation Sequencing Ngs ◽

Generation Sequencing

Next-generation sequencing (NGS) platforms have been adapted to generate genome-wide maps and sequence context of binding and cleavage of DNA topoisomerases (topos). Continuous refinements of these techniques have resulted in the acquisition of data with unprecedented depth and resolution, which has shed new light on in vivo topo behavior. Topos regulate DNA topology through the formation of reversible single- or double-stranded DNA breaks. Topo activity is critical for DNA metabolism in general, and in particular to support transcription and replication. However, the binding and activity of topos over the genome in vivo was difficult to study until the advent of NGS. Over and above traditional chromatin immunoprecipitation (ChIP)-seq approaches that probe protein binding, the unique formation of covalent protein–DNA linkages associated with DNA cleavage by topos affords the ability to probe cleavage and, by extension, activity over the genome. NGS platforms have facilitated genome-wide studies mapping the behavior of topos in vivo, how the behavior varies among species and how inhibitors affect cleavage. Many NGS approaches achieve nucleotide resolution of topo binding and cleavage sites, imparting an extent of information not previously attainable. We review the development of NGS approaches to probe topo interactions over the genome in vivo and highlight general conclusions and quandaries that have arisen from this rapidly advancing field of topoisomerase research.

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A customized scaffolds approach for the detection and phasing of complex variants by next-generation sequencing

Scientific Reports ◽

10.1038/s41598-020-71471-3 ◽

2020 ◽

Vol 10 (1) ◽

Author(s):

Qiandong Zeng ◽

Natalia T. Leach ◽

Zhaoqing Zhou ◽

Hui Zhu ◽

Jean A. Smith ◽

...

Keyword(s):

Next Generation Sequencing ◽

Variant Calling ◽

Causative Mutation ◽

Next Generation ◽

Single Nucleotide ◽

Highly Sensitive ◽

Highly Sensitive Detection ◽

Next Generation Sequencing Ngs ◽

Reference Bias ◽

Generation Sequencing

Abstract Next-generation sequencing (NGS) is widely used in genetic testing for the highly sensitive detection of single nucleotide changes and small insertions or deletions. However, detection and phasing of structural variants, especially in repetitive or homologous regions, can be problematic due to uneven read coverage or genome reference bias, resulting in false calls. To circumvent this challenge, a computational approach utilizing customized scaffolds as supplementary reference sequences for read alignment was developed, and its effectiveness demonstrated with two CBS gene variants: NM_000071.2:c.833T>C and NM_000071.2:c.[833T>C; 844_845ins68]. Variant c.833T>C is a known causative mutation for homocystinuria, but is not pathogenic when in cis with the insertion, c.844_845ins68, because of alternative splicing. Using simulated reads, the custom scaffolds method resolved all possible combinations with 100% accuracy and, based on > 60,000 clinical specimens, exceeded the performance of current approaches that only align reads to GRCh37/hg19 for the detection of c.833T>C alone or in cis with c.844_845ins68. Furthermore, analysis of two 1000 Genomes Project trios revealed that the c.[833T>C; 844_845ins68] complex variant had previously been undetected in these datasets, likely due to the alignment method used. This approach can be configured for existing workflows to detect other challenging and potentially underrepresented variants, thereby augmenting accurate variant calling in clinical NGS testing.

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A support vector machine for identification of single-nucleotide polymorphisms from next-generation sequencing data

Bioinformatics ◽

10.1093/bioinformatics/btt172 ◽

2013 ◽

Vol 29 (11) ◽

pp. 1361-1366 ◽

Cited By ~ 26

Author(s):

B. D. O'Fallon ◽

W. Wooderchak-Donahue ◽

D. K. Crockett

Keyword(s):

Support Vector Machine ◽

Next Generation Sequencing ◽

Single Nucleotide Polymorphisms ◽

Next Generation Sequencing Data ◽

Support Vector ◽

Nucleotide Polymorphisms ◽

Next Generation ◽

Sequencing Data ◽

Single Nucleotide ◽

Generation Sequencing

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Simultaneous human platelet antigen genotyping and detection of novel single nucleotide polymorphisms by targeted next-generation sequencing

Transfusion ◽

10.1111/trf.14092 ◽

2017 ◽

Vol 57 (6) ◽

pp. 1497-1504 ◽

Cited By ~ 4

Author(s):

Sue Davey ◽

Cristina Navarrete ◽

Colin Brown

Keyword(s):

Next Generation Sequencing ◽

Single Nucleotide Polymorphisms ◽

Human Platelet ◽

Nucleotide Polymorphisms ◽

Next Generation ◽

Single Nucleotide ◽

Targeted Next Generation Sequencing ◽

Platelet Antigen ◽

Human Platelet Antigen ◽

Generation Sequencing

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Abstract 4843: A next generation sequencing (NGS) genome wide copy number variation (CNV) assay for comparison of circulating tumor cell (CTC) heterogeneity

10.1158/1538-7445.am2015-4843 ◽

2015 ◽

Author(s):

Stephanie Green ◽

Mark Landers ◽

Jessica Louw ◽

Adam Jendrisak ◽

Ryan Dittamore ◽

...

Keyword(s):

Next Generation Sequencing ◽

Copy Number Variation ◽

Tumor Cell ◽

Copy Number ◽

Circulating Tumor Cell ◽

Next Generation ◽

Genome Wide ◽

Number Variation ◽

Next Generation Sequencing Ngs ◽

Generation Sequencing

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Next-Generation Sequencing Approaches in Genome-Wide Discovery of Single Nucleotide Polymorphism Markers Associated with Pungency and Disease Resistance in Pepper

BioMed Research International ◽

10.1155/2018/5646213 ◽

2018 ◽

Vol 2018 ◽

pp. 1-7 ◽

Cited By ~ 3

Author(s):

Abinaya Manivannan ◽

Jin-Hee Kim ◽

Eun-Young Yang ◽

Yul-Kyun Ahn ◽

Eun-Su Lee ◽

...

Keyword(s):

Single Nucleotide Polymorphism ◽

Disease Resistance ◽

Next Generation Sequencing ◽

Molecular Mechanisms ◽

Next Generation ◽

Nucleotide Polymorphism ◽

Single Nucleotide ◽

Genome Wide ◽

The Impact ◽

Generation Sequencing

Pepper is an economically important horticultural plant that has been widely used for its pungency and spicy taste in worldwide cuisines. Therefore, the domestication of pepper has been carried out since antiquity. Owing to meet the growing demand for pepper with high quality, organoleptic property, nutraceutical contents, and disease tolerance, genomics assisted breeding techniques can be incorporated to develop novel pepper varieties with desired traits. The application of next-generation sequencing (NGS) approaches has reformed the plant breeding technology especially in the area of molecular marker assisted breeding. The availability of genomic information aids in the deeper understanding of several molecular mechanisms behind the vital physiological processes. In addition, the NGS methods facilitate the genome-wide discovery of DNA based markers linked to key genes involved in important biological phenomenon. Among the molecular markers, single nucleotide polymorphism (SNP) indulges various benefits in comparison with other existing DNA based markers. The present review concentrates on the impact of NGS approaches in the discovery of useful SNP markers associated with pungency and disease resistance in pepper. The information provided in the current endeavor can be utilized for the betterment of pepper breeding in future.

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