scholarly journals Next-generation polyploid phylogenetics: rapid resolution of hybrid polyploid complexes using PacBio single-molecule sequencing

2016 ◽  
Vol 213 (1) ◽  
pp. 413-429 ◽  
Author(s):  
Carl J. Rothfels ◽  
Kathleen M. Pryer ◽  
Fay-Wei Li
Keyword(s):  
Single Molecule ◽  
Next Generation ◽  
Single Molecule Sequencing ◽  
Rapid Resolution

scholarly journals HySA: A Hybrid Structural variant Assembly approach using next generation and single-molecule sequencing technologies

2016 ◽  
Author(s):  
Xian Fan ◽  
Mark Chaisson ◽  
Luay Nakhleh ◽  
Ken Chen
Keyword(s):  
Human Genome ◽  
Single Molecule ◽  
Clustering Algorithm ◽  
Hydatidiform Mole ◽  
Cost Effective ◽  
Next Generation ◽  
Structural Variations ◽  
Single Molecule Sequencing ◽  
Structural Variant ◽  
Sequencing Technologies

AbstractAchieving complete, accurate and cost-effective assembly of human genome is of great importance for realizing the promises of precision medicine. The abundance of repeats and genetic variations in human genome and the limitations of existing sequencing technologies call for the development of novel assembly methods that could leverage the complementary strengths of multiple technologies.We propose a Hybrid Structural variant Assembly (HySA) approach that integrates sequencing reads from next generation sequencing (NGS) and single-molecule sequencing (SMS) technologies to accurately assemble and detect structural variations (SV) in human genome. By identifying homologous SV-containing reads from different technologies through a bipartite-graph-based clustering algorithm, our approach turns a whole genome assembly problem into a set of independent SV assembly problems, each of which can be effectively solved to enhance assembly of structurally altered regions in human genome.In testing our approach using data generated from a haploid hydatidiform mole genome (CHM1) and a diploid human genome (NA12878), we found that our approach substantially improved the detection of many types of SVs, particularly novel large insertions, small INDELs (10-50bp) and short tandem repeat expansions and contractions over existing approaches with a low false discovery rate. Our work highlights the strengths and limitations of current approaches and provides an effective solution for extending the power of existing sequencing technologies for SV discovery.

scholarly journals Accurate CNV identification from only a few cells with low GC bias in a single-molecule sequencing platform

2020 ◽  
Author(s):  
Liang Hu ◽  
Qunting Lin ◽  
Pingyuan Xie ◽  
Lidong Zeng ◽  
Lichun Liu ◽  
...  
Keyword(s):  
Next Generation Sequencing ◽  
Single Molecule ◽  
Technical Problem ◽  
Next Generation ◽  
Single Molecule Sequencing ◽  
Sequencing Platform ◽  
Transposon Insertion ◽  
Number Variation ◽  
Gc Bias ◽  
Generation Sequencing

ABSTRACTA technical problem of characterizing copy number variation of several cells with next-generation sequencing is the whole genome amplification induced bias. The result of CNVs and mosaicism detection is affected by the GC bias. Here, we report a rapid non-WGA sample preparation strategy for a single-molecule sequencing platform GenoCare1600. This approach, combined with a single-molecule sequencing platform that avoids the use of WGA and bridge PCR processes, can provide higher reliability with its lower GC bias. By combining our optimized Tn5-based transposon insertion approach with GenoCare, we successfully detected CNVs as small as 1.29M and mosaicism as small as 20%, which is consistent with next-generation sequencing (NGS) data. Moreover, our GenoCare-TTI protocol showed less GC bias and less Mad of Diff. These results suggest that the optimized TTI approach, together with the GenoCare1600 sequencing platform, is a promising option for CNV characterization from maybe one single cell.

scholarly journals Next-Generation Sequencing: From Basic Research to Diagnostics

2009 ◽  
Vol 55 (4) ◽  
pp. 641-658 ◽  
Author(s):  
Karl V Voelkerding ◽  
Shale A Dames ◽  
Jacob D Durtschi
Keyword(s):  
Next Generation Sequencing ◽  
Dna Sequencing ◽  
Single Molecule ◽  
Basic Research ◽  
Clinical Diagnostics ◽  
Massively Parallel ◽  
Next Generation ◽  
New Paradigm ◽  
The Impact ◽  
Generation Sequencing

Abstract Background: For the past 30 years, the Sanger method has been the dominant approach and gold standard for DNA sequencing. The commercial launch of the first massively parallel pyrosequencing platform in 2005 ushered in the new era of high-throughput genomic analysis now referred to as next-generation sequencing (NGS). Content: This review describes fundamental principles of commercially available NGS platforms. Although the platforms differ in their engineering configurations and sequencing chemistries, they share a technical paradigm in that sequencing of spatially separated, clonally amplified DNA templates or single DNA molecules is performed in a flow cell in a massively parallel manner. Through iterative cycles of polymerase-mediated nucleotide extensions or, in one approach, through successive oligonucleotide ligations, sequence outputs in the range of hundreds of megabases to gigabases are now obtained routinely. Highlighted in this review are the impact of NGS on basic research, bioinformatics considerations, and translation of this technology into clinical diagnostics. Also presented is a view into future technologies, including real-time single-molecule DNA sequencing and nanopore-based sequencing. Summary: In the relatively short time frame since 2005, NGS has fundamentally altered genomics research and allowed investigators to conduct experiments that were previously not technically feasible or affordable. The various technologies that constitute this new paradigm continue to evolve, and further improvements in technology robustness and process streamlining will pave the path for translation into clinical diagnostics.

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