July 19, 2019  |  

Multiplexed highly-accurate DNA sequencing of closely-related HIV-1 variants using continuous long reads from single molecule, real-time sequencing.

Authors: Dilernia, Dario A and Chien, Jung-Ting and Monaco, Daniela C and Brown, Michael P S and Ende, Zachary and Deymier, Martin J and Yue, Ling and Paxinos, Ellen E and Allen, Susan and Tirado-Ramos, Alfredo and Hunter, Eric

Single Molecule, Real-Time (SMRT(®)) Sequencing (Pacific Biosciences, Menlo Park, CA, USA) provides the longest continuous DNA sequencing reads currently available. However, the relatively high error rate in the raw read data requires novel analysis methods to deconvolute sequences derived from complex samples. Here, we present a workflow of novel computer algorithms able to reconstruct viral variant genomes present in mixtures with an accuracy of >QV50. This approach relies exclusively on Continuous Long Reads (CLR), which are the raw reads generated during SMRT Sequencing. We successfully implement this workflow for simultaneous sequencing of mixtures containing up to forty different >9 kb HIV-1 full genomes. This was achieved using a single SMRT Cell for each mixture and desktop computing power. This novel approach opens the possibility of solving complex sequencing tasks that currently lack a solution. © The Author(s) 2015. Published by Oxford University Press on behalf of Nucleic Acids Research.

Journal: Nucleic acids research
DOI: 10.1093/nar/gkv630
Year: 2015

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