![]() ![]() Protocol from the Grubaugh lab at Yale (gdoc).Probes can be ordered from Twist Bioscience we have used the protocol for Twist custom panels with slight modifications for low input Nextera XT libraries.Ī number of different laboratories have implemented derivatives of the ARTIC amplicon scheme on Illumina. It was initially released in January, 2020 and most recently updated in March, 2020. The probe set is available as V-Respiratory on the probe designs page of the CATCH repository. The Sabeti lab, at the Broad Institute, released a probe set for comprehensive whole-genome capture of SARS-CoV-2 and respiratory-related viruses (human-infecting coronaviruses, HRSV, HMPV, HPIVs, Human mastadenovirus A-G, Enterovirus A-E, Rhinovirus A/B/C, influenza A/B/C). Illumina's technical note on sequencing coronavirus samples using a comprehensive metagenomic sequencing approach was one of the earlier protocols released for SARS-CoV-2, and remains an effective option for shotgun sequencing.ĭ) SARS-CoV-2 and related virus sequencing with capture enrichment (Broad Institute) ![]() The protocol is available on protocols.io.Ĭ) Illumina Shotgun Metagenomics Sequencing Protocol The NIAID laboratory team in Cambodia, in collaboration with UCSF, CZBioHub and IPC, has released a metagenomic sequencing with spiked primer enrichment (MSSPE) protocol for SARS-CoV-2. Illumina's Research and Development group has recently developed and validated a custom, research use only (RUO) enrichment sequencing strategy based on their Nextera Flex chemistry.ī) SARS-CoV-2 Enrichment Sequencing by Spiked Primer MSSPE (NIAID/UCSF/CZBioHub) ILLUMINA a) Illumina Nextera Flex Enrichment Sequencing Protocol The Victorian Infectious Diseases Reference Laboratory ( VIDRL) at the Peter Doherty Institute for Infection and Immunity released two protocols for the ONT MinION, which they successfully used to sequence early Australian SARS-CoV-2 samples.Ģ. b) Doherty Institute VIDRL Sequencing Protocols and the ARTIC Network team describes a greatly streamlined workflow for ARTIC-Nanopore sequencing of SARS-CoV-2, including functional multiplexing library construction up to 96 samples. Integrated bioinformatics (RAMPART) - documentation below under bioinformatics methods. Sequencing protocol / Single sample sequencing protocolĪRTIC on Illumina - Complete Walk-through - Grubaugh/Andersen/Loman labs This protocol was developed and released by the fine folks at ARTIC Network, and was subsequently refined based on comments from Itokawa et al, which identified potential issues and proposed an alternate L18 primer. a) ARTIC Network nCoV-2019 Sequencing Protocol Even so, most of these protocols should scale to larger ONT instruments without significant modifications. The following sequencing protocols, checklists and job-aids are primarily designed for the Oxford Nanopore MinION, and have been kindly shared by research groups throughout the world (please see individual protocols for attribution and citing purposes). Use of trade names is for identification only and does not imply endorsement by the Centers for Disease Control and Prevention or by the U.S. The findings and conclusions in this document and the attendant repository are those of the author and do not necessarily represent the official position of the Centers for Disease Control and Prevention. Submitting to Public Sequence Repositories.Bioinformatic Tools, Scripts and Workflows.In the meantime, I'll continue to add and mirror useful resources here as they become available. Please feel free to suggest additions, edits, clarifications and corrections - either by posting an issue, filing a pull request or by contacting me directly by email or twitter. It has incorporated collaborative edits from more than two dozen submitters throughout the world, and is actively maintained by Michael Weigand and the CDC TOAST Team. This collection was originally established, maintained and curated by Duncan MacCannell from the Office of Advanced Molecular Detection ( AMD) at the Centers for Disease Control and Prevention ( CDC). While some of the resources here are directed specifically to US state and local public health laboratories in support of diagnostic testing, sequencing and response, we hope that this is a useful resource for the global laboratory community, as we respond to this pandemic threat. This is admittedly a limited first draft, but will continued to collate useful information as additional protocols, tools, and resources are added, and as best practices are identified. This document repository is meant to serve as the start of a crowd-sourced collection of information, documentation, protocols and other resources for public health laboratories intending to sequence SARS-CoV-2 coronavirus samples in the coming weeks. ![]()
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