scholarly journals Ultrasensitive amplicon barcoding for next-generation sequencing facilitating sequence error and amplification-bias correction

2020 ◽  
Vol 10 (1) ◽  
Author(s):  
Ibrahim Ahmed ◽  
Felicia A. Tucci ◽  
Aure Aflalo ◽  
Kenneth G. C. Smith ◽  
Rachael J. M. Bashford-Rogers
Keyword(s):  
Next Generation Sequencing ◽  
Bias Correction ◽  
Tumor Heterogeneity ◽  
Pcr Amplification ◽  
Genetic Studies ◽  
Immune Receptor ◽  
Amplification Bias ◽  
Sequencing Errors ◽  
Sequence Error ◽  
Generation Sequencing

Abstract The ability to accurately characterize DNA variant proportions using PCR amplification is key to many genetic studies, including studying tumor heterogeneity, 16S microbiome, viral and immune receptor sequencing. We develop a novel generalizable ultrasensitive amplicon barcoding approach that significantly reduces the inflation/deflation of DNA variant proportions due to PCR amplification biases and sequencing errors. This method was applied to immune receptor sequencing, where it significantly improves the quality and estimation of diversity of the resulting library.

scholarly journals Author Correction: Ultrasensitive amplicon barcoding for next-generation sequencing facilitating sequence error and amplification-bias correction

2020 ◽  
Vol 10 (1) ◽  
Author(s):  
Ibrahim Ahmed ◽  
Felicia A. Tucci ◽  
Aure Aflalo ◽  
Kenneth G. C. Smith ◽  
Rachael J. M. Bashford-Rogers
Keyword(s):  
Next Generation Sequencing ◽  
Bias Correction ◽  
Next Generation ◽  
Amplification Bias ◽  
Sequence Error ◽  
Generation Sequencing

An amendment to this paper has been published and can be accessed via a link at the top of the paper.

scholarly journals Next-Generation Sequencing in Human Genetic Studies: Genome Technologies and Applications to Human Genetic Studies

2018 ◽  
Vol 83 (3) ◽  
pp. 105-106
Author(s):  
Junwen Wang ◽  
Kai Wang ◽  
Xiaoming Liu ◽  
Pak Sham ◽  
Zhongming Zhao
Keyword(s):  
Next Generation Sequencing ◽  
Next Generation ◽  
Genetic Studies ◽  
Generation Sequencing

Quantitative evaluation of bias in PCR amplification and next-generation sequencing derived from metabarcoding samples

2015 ◽  
Vol 407 (7) ◽  
pp. 1841-1848 ◽  
Author(s):  
Marta Pawluczyk ◽  
Julia Weiss ◽  
Matthew G. Links ◽  
Mikel Egaña Aranguren ◽  
Mark D. Wilkinson ◽  
...  
Keyword(s):  
Next Generation Sequencing ◽  
Quantitative Evaluation ◽  
Pcr Amplification ◽  
Next Generation ◽  
Generation Sequencing

scholarly journals Next Generation Sequencing of Pooled Samples: Guideline for Variants’ Filtering

2016 ◽  
Vol 6 (1) ◽  
Author(s):  
Santosh Anand ◽  
Eleonora Mangano ◽  
Nadia Barizzone ◽  
Roberta Bordoni ◽  
Melissa Sorosina ◽  
...  
Keyword(s):  
Next Generation Sequencing ◽  
Low Frequency ◽  
Next Generation ◽  
Sequencing Data ◽  
Sequencing Errors ◽  
Effective Option ◽  
Sequencing Experiment ◽  
Kolmogorov Smirnov ◽  
Next Generation Sequencing Ngs ◽  
Generation Sequencing

Abstract Sequencing large number of individuals, which is often needed for population genetics studies, is still economically challenging despite falling costs of Next Generation Sequencing (NGS). Pool-seq is an alternative cost- and time-effective option in which DNA from several individuals is pooled for sequencing. However, pooling of DNA creates new problems and challenges for accurate variant call and allele frequency (AF) estimation. In particular, sequencing errors confound with the alleles present at low frequency in the pools possibly giving rise to false positive variants. We sequenced 996 individuals in 83 pools (12 individuals/pool) in a targeted re-sequencing experiment. We show that Pool-seq AFs are robust and reliable by comparing them with public variant databases and in-house SNP-genotyping data of individual subjects of pools. Furthermore, we propose a simple filtering guideline for the removal of spurious variants based on the Kolmogorov-Smirnov statistical test. We experimentally validated our filters by comparing Pool-seq to individual sequencing data showing that the filters remove most of the false variants while retaining majority of true variants. The proposed guideline is fairly generic in nature and could be easily applied in other Pool-seq experiments.

scholarly journals Microsatellite loci discovery from next-generation sequencing data and loci characterization in the epizoic barnacleChelonibia testudinaria(Linnaeus, 1758)

PeerJ ◽  
2016 ◽  
Vol 4 ◽  
pp. e2019 ◽  
Author(s):  
Christine Ewers-Saucedo ◽  
John D. Zardus ◽  
John P. Wares
Keyword(s):  
Next Generation Sequencing ◽  
Microsatellite Markers ◽  
Microsatellite Loci ◽  
Next Generation Sequencing Data ◽  
Model Organisms ◽  
Next Generation ◽  
Sequencing Data ◽  
Genetic Studies ◽  
Evolutionary Features ◽  
Generation Sequencing

Microsatellite markers remain an important tool for ecological and evolutionary research, but are unavailable for many non-model organisms. One such organism with rare ecological and evolutionary features is the epizoic barnacleChelonibia testudinaria(Linnaeus, 1758).Chelonibia testudinariaappears to be a host generalist, and has an unusual sexual system, androdioecy. Genetic studies on host specificity and mating behavior are impeded by the lack of fine-scale, highly variable markers, such as microsatellite markers. In the present study, we discovered thousands of new microsatellite loci from next-generation sequencing data, and characterized 12 loci thoroughly. We conclude that 11 of these loci will be useful markers in future ecological and evolutionary studies onC. testudinaria.

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