scholarly journals De Novo Assembly-Based Analysis of RPGR Exon ORF15 in an Indigenous African Cohort Overcomes Limitations of a Standard Next-Generation Sequencing (NGS) Data Analysis Pipeline

Genes ◽  
2020 ◽  
Vol 11 (7) ◽  
pp. 800
Author(s):  
Jordi Maggi ◽  
Lisa Roberts ◽  
Samuel Koller ◽  
George Rebello ◽  
Wolfgang Berger ◽  
...  
Keyword(s):  
Data Analysis ◽  
Next Generation Sequencing ◽  
De Novo Assembly ◽  
De Novo ◽  
African Ancestry ◽  
Secondary Analysis ◽  
Next Generation ◽  
Analysis Pipeline ◽  
Next Generation Sequencing Ngs ◽  
Generation Sequencing

RPGR exon ORF15 variants are one of the most frequent causes for inherited retinal disorders (IRDs), in particular retinitis pigmentosa. The low sequence complexity of this mutation hotspot makes it prone to indels and challenging for sequence data analysis. Whole-exome sequencing generally fails to provide adequate coverage in this region. Therefore, complementary methods are needed to avoid false positives as well as negative results. In this study, next-generation sequencing (NGS) was used to sequence long-range PCR amplicons for an IRD cohort of African ancestry. By developing a novel secondary analysis pipeline based on de novo assembly, we were able to avoid the miscalling of variants generated by standard NGS analysis tools. We identified pathogenic variants in 11 patients (13% of the cohort), two of which have not been reported previously. We provide a novel and alternative end-to-end secondary analysis pipeline for targeted NGS of ORF15 that is less prone to false positive and negative variant calls.

scholarly journals Compression of next-generation sequencing reads aided by highly efficient de novo assembly

2012 ◽  
Vol 40 (22) ◽  
pp. e171-e171 ◽  
Author(s):  
Daniel C. Jones ◽  
Walter L. Ruzzo ◽  
Xinxia Peng ◽  
Michael G. Katze
Keyword(s):  
Next Generation Sequencing ◽  
De Novo Assembly ◽  
De Novo ◽  
Next Generation ◽  
Highly Efficient ◽  
Generation Sequencing

scholarly journals Applications and data analysis of next-generation sequencing

2013 ◽  
Vol 37 (6) ◽  
Author(s):  
Ina Vogl ◽  
Anna Benet-Pagès ◽  
Sebastian H. Eck ◽  
Marius Kuhn ◽  
Sebastian Vosberg ◽  
...  
Keyword(s):  
Data Analysis ◽  
Next Generation Sequencing ◽  
Next Generation ◽  
High Expectations ◽  
Sequencing Platform ◽  
Software Products ◽  
High Throughput Method ◽  
Next Generation Sequencing Ngs ◽  
Generation Sequencing ◽  
Enrichment Methods

Abstract:Over the past 6 years, next-generation sequencing (NGS) has been established as a valuable high-throughput method for research in molecular genetics and has successfully been employed in the identification of rare and common genetic variations. Although the high expectations regarding the discovery of new diagnostic targets and an overall reduction of cost have been achieved, technological challenges in instrument handling, robustness of the chemistry, and data analysis need to be overcome. Each workflow and sequencing platform have their particular problems and caveats, which need to be addressed. Regarding NGS, there is a variety of different enrichment methods, sequencing devices, or technologies as well as a multitude of analyzing software products available. In this manuscript, the authors focus on challenges in data analysis when employing different target enrichment methods and the best applications for each of them.

scholarly journals An ensemble strategy that significantly improves de novo assembly of microbial genomes from metagenomic next-generation sequencing data

2015 ◽  
Vol 43 (7) ◽  
pp. e46-e46 ◽  
Author(s):  
Xutao Deng ◽  
Samia N. Naccache ◽  
Terry Ng ◽  
Scot Federman ◽  
Linlin Li ◽  
...  
Keyword(s):  
Next Generation Sequencing ◽  
De Novo Assembly ◽  
De Novo ◽  
Next Generation Sequencing Data ◽  
De Bruijn Graph ◽  
Next Generation ◽  
Sequencing Data ◽  
Short Reads ◽  
Ensemble Strategy ◽  
Generation Sequencing

Abstract Next-generation sequencing (NGS) approaches rapidly produce millions to billions of short reads, which allow pathogen detection and discovery in human clinical, animal and environmental samples. A major limitation of sequence homology-based identification for highly divergent microorganisms is the short length of reads generated by most highly parallel sequencing technologies. Short reads require a high level of sequence similarities to annotated genes to confidently predict gene function or homology. Such recognition of highly divergent homologues can be improved by reference-free (de novo) assembly of short overlapping sequence reads into larger contigs. We describe an ensemble strategy that integrates the sequential use of various de Bruijn graph and overlap-layout-consensus assemblers with a novel partitioned sub-assembly approach. We also proposed new quality metrics that are suitable for evaluating metagenome de novo assembly. We demonstrate that this new ensemble strategy tested using in silico spike-in, clinical and environmental NGS datasets achieved significantly better contigs than current approaches.

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