Multiplex PCR method for MinION and Illumina sequencing of Zika and other virus genomes directly from clinical samples
Joshua Quick,Nathan D. Grubaugh,Steven T. Pullan,Ingra Morales Claro,Andrew D. Smith,Karthik Gangavarapu,Glenn Oliveira,Refugio Robles-Sikisaka,Thomas F. Rogers,Thomas F. Rogers,Nathan Beutler,Dennis R. Burton,Lia Laura Lewis-Ximenez,Jaqueline Goes de Jesus,Marta Giovanetti,Marta Giovanetti,Sarah C. Hill,Allison Black,Allison Black,Trevor Bedford,Miles W. Carroll,Márcio Roberto Teixeira Nunes,Luiz Carlos Junior Alcantara,Ester Cerdeira Sabino,Sally A. Baylis,Nuno R. Faria,Matthew Loose,Jared T. Simpson,Oliver G. Pybus,Kristian G. Andersen,Kristian G. Andersen,Nicholas J. Loman +31 more
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TLDR
A protocol for generating coding-sequence-complete genomes, comprising an online primer design tool, a novel multiplex PCR enrichment protocol, optimized library preparation methods for the portable MinION sequencer and the Illumina range of instruments, and a bioinformatics pipeline for generating consensus sequences.Abstract:
Genome sequencing has become a powerful tool for studying emerging infectious diseases; however, genome sequencing directly from clinical samples (ie, without isolation and culture) remains challenging for viruses such as Zika, for which metagenomic sequencing methods may generate insufficient numbers of viral reads Here we present a protocol for generating coding-sequence-complete genomes, comprising an online primer design tool, a novel multiplex PCR enrichment protocol, optimized library preparation methods for the portable MinION sequencer (Oxford Nanopore Technologies) and the Illumina range of instruments, and a bioinformatics pipeline for generating consensus sequences The MinION protocol does not require an Internet connection for analysis, making it suitable for field applications with limited connectivity Our method relies on multiplex PCR for targeted enrichment of viral genomes from samples containing as few as 50 genome copies per reaction Viral consensus sequences can be achieved in 1-2 d by starting with clinical samples and following a simple laboratory workflow This method has been successfully used by several groups studying Zika virus evolution and is facilitating an understanding of the spread of the virus in the Americas The protocol can be used to sequence other viral genomes using the online Primal Scheme primer designer software It is suitable for sequencing either RNA or DNA viruses in the field during outbreaks or as an inexpensive, convenient method for use in the labread more
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SPAdes, a new genome assembly algorithm and its applications to single-cell sequencing ( 7th Annual SFAF Meeting, 2012)
TL;DR: SPAdes as mentioned in this paper is a new assembler for both single-cell and standard (multicell) assembly, and demonstrate that it improves on the recently released E+V-SC assembler and on popular assemblers Velvet and SoapDeNovo (for multicell data).
Integrative Genomics Viewer
James T. Robinson,Helga Thorvaldsdottir,Wendy Winckler,Mitchell Guttman,Eric S. Lander,Eric S. Lander,Gad Getz,Jill P. Mesirov +7 more
TL;DR: The sheer volume and scope of data posed by this flood of data pose a significant challenge to the development of efficient and intuitive visualization tools able to scale to very large data sets and to flexibly integrate multiple data types, including clinical data.
Journal ArticleDOI
Nanopore sequencing and assembly of a human genome with ultra-long reads
Miten Jain,Sergey Koren,Karen H. Miga,Josh Quick,Arthur C Rand,Thomas A Sasani,John R. Tyson,Andrew D Beggs,Alexander T. Dilthey,Ian T. Fiddes,Sunir Malla,Hannah Marriott,Tom Nieto,Justin O'Grady,Hugh E. Olsen,Brent S. Pedersen,Arang Rhie,Hollian Richardson,Aaron R. Quinlan,Terrance P. Snutch,Louise Tee,Benedict Paten,Adam M. Phillippy,Jared T. Simpson,Jared T. Simpson,Nicholas J. Loman,Matthew Loose +26 more
TL;DR: Ultra-long reads enabled assembly and phasing of the 4-Mb major histocompatibility complex (MHC) locus in its entirety, measurement of telomere repeat length, and closure of gaps in the reference human genome assembly GRCh38.
Journal ArticleDOI
Neuropilin-1 facilitates SARS-CoV-2 cell entry and infectivity.
Ludovico Cantuti-Castelvetri,Ludovico Cantuti-Castelvetri,Ravi Ojha,Liliana D. Pedro,Liliana D. Pedro,Minou Djannatian,Minou Djannatian,Jonas Franz,Suvi Kuivanen,Franziska van der Meer,Katri Kallio,Tuğberk Kaya,Tuğberk Kaya,Tuğberk Kaya,Maria Anastasina,Maria Anastasina,Teemu Smura,Lev Levanov,Leonora Szirovicza,Allan Tobi,Hannimari Kallio-Kokko,Pamela Österlund,Merja Joensuu,Frederic A. Meunier,Sarah J. Butcher,Sarah J. Butcher,Martin Sebastian Winkler,Brit Mollenhauer,Ari Helenius,Ozgun Gokce,Tambet Teesalu,Tambet Teesalu,Tambet Teesalu,Jussi Hepojoki,Jussi Hepojoki,Olli Vapalahti,Christine Stadelmann,Giuseppe Balistreri,Giuseppe Balistreri,Mikael Simons,Mikael Simons +40 more
TL;DR: It is found that neuropilin-1 (NRP1), known to bind furin-cleaved substrates, significantly potentiates SARS-CoV-2 infectivity, an effect blocked by a monoclonal blocking antibody against NRP1.
Journal ArticleDOI
Viral and host factors related to the clinical outcome of COVID-19.
Xiaonan Zhang,Yun Tan,Yun Ling,Gang Lu,Feng Liu,Zhigang Yi,Xiaofang Jia,Min Wu,Bisheng Shi,Shuibao Xu,Jun Chen,Wei Wang,Bing Chen,Lu Jiang,Shuting Yu,Jing Lu,Jinzeng Wang,Mingzhu Xu,Zhenghong Yuan,Qin Zhang,Xinxin Zhang,Guoping Zhao,Shengyue Wang,Sai-Juan Chen,Hongzhou Lu +24 more
TL;DR: Clinical data from 326 cases suggest that T cell depletion and cytokine bursts are associated with a worse prognosis, and genomic sequences from 112 patients with confirmed SARS-CoV-2 virus showed two clades with similar virulence and clinical outcome.
References
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Integrative genomics viewer
James T. Robinson,Helga Thorvaldsdottir,Wendy Winckler,Mitchell Guttman,Eric S. Lander,Eric S. Lander,Gad Getz,Jill P. Mesirov +7 more
TL;DR: In this article, the authors present an approach for efficient and intuitive visualization tools able to scale to very large data sets and to flexibly integrate multiple data types, including clinical data.
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