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An amplicon-based sequencing framework for accurately measuring intrahost virus diversity using PrimalSeq and iVar

by Quick, Joshua , Gangavarapu, Karthik , Brackney, Doug E. , Grubaugh, Nathan D. , Coffey, Lark L. , Main, Bradley J. , Tan, Amanda L. , Van Rompay, Koen K. A. , Loman, Nicholas J. , Michael, Scott F. , Isern, Sharon , Gurfield, Nikos , De Jesus, Jaqueline Goes , Paul, Lauren M. , Andersen, Kristian G. , Matteson, Nathaniel L. , Grewal, Saran

in Accuracy / Amplicon sequencing / Animal Genetics and Genomics / Bias / Bioinformatics / Biomedical and Life Sciences / Computer applications / Evolution / Evolutionary Biology / Genetic diversity / Genetic Variation / genome / Genomes / Genomics - methods / Human Genetics / Infections / Intrahost evolution / Laboratories / Life Sciences / Method / Microbial Genetics and Genomics / Mosquitoes / nanopores / Plant Genetics and Genomics / Population / Protocol / Sequence Analysis, RNA / SNP calling / Viral sequencing / viruses / West Nile / West Nile virus / West Nile virus - genetics / Zika / Zika virus / Zika Virus - genetics

2019

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An amplicon-based sequencing framework for accurately measuring intrahost virus diversity using PrimalSeq and iVar

2019

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An amplicon-based sequencing framework for accurately measuring intrahost virus diversity using PrimalSeq and iVar

2019

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Journal Article

An amplicon-based sequencing framework for accurately measuring intrahost virus diversity using PrimalSeq and iVar

Quick, Joshua,

Gangavarapu, Karthik,

Brackney, Doug E.,

Grubaugh, Nathan D.,

Coffey, Lark L.,

Main, Bradley J.,

Tan, Amanda L.,

Van Rompay, Koen K. A.,

Loman, Nicholas J.,

Michael, Scott F.,

Isern, Sharon,

Gurfield, Nikos,

De Jesus, Jaqueline Goes,

Paul, Lauren M.,

Andersen, Kristian G.,

Matteson, Nathaniel L.,

Grewal, Saran

2019

Overview

How viruses evolve within hosts can dictate infection outcomes; however, reconstructing this process is challenging. We evaluate our multiplexed amplicon approach, PrimalSeq, to demonstrate how virus concentration, sequencing coverage, primer mismatches, and replicates influence the accuracy of measuring intrahost virus diversity. We develop an experimental protocol and computational tool, iVar, for using PrimalSeq to measure virus diversity using Illumina and compare the results to Oxford Nanopore sequencing. We demonstrate the utility of PrimalSeq by measuring Zika and West Nile virus diversity from varied sample types and show that the accumulation of genetic diversity is influenced by experimental and biological systems.

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Publisher

BioMed Central,Springer Nature B.V,BMC

Subject

Accuracy

/ Amplicon sequencing

/ Animal Genetics and Genomics

/ Bias

/ Bioinformatics

/ Biomedical and Life Sciences

/ Computer applications