Mar 21, 2024

Public workspaceGenomeTrakr WGS Protocol Collection and Workflow for MiSeq V.3

DOI
dx.doi.org/10.17504/protocols.io.3byl4bwyjvo5/v3
  • 1US Food and Drug Administration
Tina Lusk Pfefer
Open access
DOI: dx.doi.org/10.17504/protocols.io.3byl4bwyjvo5/v3
Protocol Citation: Tina Lusk Pfefer, Julie Haendiges, Maria Balkey, Candace Hope Bias, Ruth Timme 2024. GenomeTrakr WGS Protocol Collection and Workflow for MiSeq. protocols.io https://dx.doi.org/10.17504/protocols.io.3byl4bwyjvo5/v3Version created by Tina Lusk Pfefer
License: This is an open access protocol distributed under the terms of the Creative Commons Attribution License,  which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited
Protocol status: Working
We use this protocol and it's working
Created: March 19, 2024
Last Modified: March 21, 2024
Protocol Integer ID: 96944
Keywords: GenomeTrakr, whole genome sequencing, enteric pathogens, surveillance, MiSeq
Abstract
Here we have created a collection of all the protocols used for WGS using the MiSeq, in order, from sample extraction to NCBI submission.

This collection has three sections:

  • Section 1: WGS Wet lab workflow for Illumina MiSeq
  • Section 2: Dry lab workflow for sequence QC and NCBI submission - Direct Submission
  • Section 3: Dry lab workflow for sequence QC and NCBI submission - PulseNet labs

Version history:
V3:
  • Links to protocols updated with latest version,
  • Added new protocol: DNA Extraction from Inoculated Broth for WGS using the Maxwell RSC Cultured Cells DNA Kit,
  • Added new protocol: Querying for Bacterial Pathogen Genomic Data at NCBI.
WGS Wet Lab Workflow for Illumina MiSeq
WGS Wet Lab Workflow for Illumina MiSeq
DNA Extraction - Manual
Protocol
Manual DNA Extraction using Qiagen DNeasy Blood and Tissue Kit
NAME

Manual DNA Extraction using Qiagen DNeasy Blood and Tissue Kit

CREATED BY
Julie Haendiges

DNA Extraction - Automated


Protocol
DNA Extraction from Inoculated Broth for WGS using the Maxwell RSC Cultured Cells DNA Kit
NAME
DNA Extraction from Inoculated Broth for WGS using the Maxwell RSC Cultured Cells DNA Kit
CREATED BY
Julie Haendiges

DNA Quantification
Protocol
DNA Quantification using the Qubit Fluorometer
NAME

DNA Quantification using the Qubit Fluorometer

CREATED BY
Julie Haendiges

Library Preparation

Protocol
Illumina DNA Prep (M) Tagmentation Library Preparation for use on an Illumina MiSeq Sequencer
NAME
Illumina DNA Prep (M) Tagmentation Library Preparation for use on an Illumina MiSeq Sequencer
CREATED BY
Julie Haendiges

Sequencing
Protocol
Procedure for Operation and Maintenance of the Illumina MiSeq for Whole Genome Sequencing
NAME

Procedure for Operation and Maintenance of the Illumina MiSeq for Whole Genome Sequencing

CREATED BY
Julie Haendiges

Dry Lab Workflow for Sequence QC and NCBI Submission - Direct Submission
Dry Lab Workflow for Sequence QC and NCBI Submission - Direct Submission
The following protocols are also included in a Springer Methods book chapter collection: Utilizing the Public GenomeTrakr Database for Foodborne Pathogen Traceback (protocols.io)

Check sequence quality:
Protocol
Quality control assessment for microbial genomes: GalaxyTrakr MicroRunQC workflow
NAME
Quality control assessment for microbial genomes: GalaxyTrakr MicroRunQC workflow
CREATED BY
Ruth Timme

Populate BioSample AND SRA metadata templates:
Protocol
Guidance for populating and validating GenomeTrakr metadata templates (BioSample and SRA)
NAME
Guidance for populating and validating GenomeTrakr metadata templates (BioSample and SRA)
CREATED BY
Ruth Timme

Submit sequence and metadata to NCBI:
Protocol
NCBI submission protocol for microbial pathogen surveillance
NAME
NCBI submission protocol for microbial pathogen surveillance
CREATED BY
Ruth Timme

Querying data at NCBI
Protocol
Querying for Bacterial Pathogen Genomic Data at NCBI
NAME
Querying for Bacterial Pathogen Genomic Data at NCBI
CREATED BY
Maria Balkey

Update, retract, or replace your submissions at NCBI:
Protocol
NCBI Bacterial Pathogen Data Curation Protocol: SOP for Editing GenomeTrakr Submissions
NAME
NCBI Bacterial Pathogen Data Curation Protocol: SOP for Editing GenomeTrakr Submissions
CREATED BY
Ruth Timme

Dry Lab Workflow for Sequence QC and NCBI Submission - PulseNet Labs
Dry Lab Workflow for Sequence QC and NCBI Submission - PulseNet Labs
Check sequence quality:

Follow the CDC's guidance for assessing QC in BioNumerics
Populate NCBI template:

***NOTE: For NCBI submission for fields not included in the BioNumerics Template, please remember to include the name of the laboratory sequencing the isolates and the surveillance effort name in the sequenced_by and project_name fields as described in Step 5 of this protocol.

Protocol
Populating NCBI template for submissions using BioNumerics
NAME
Populating NCBI template for submissions using BioNumerics
CREATED BY
Maria Balkey

Submit sequence and metadata to NCBI:

Follow the CDC's guidance for NCBI submissions through BioNumerics.
Query data at NCBI

Go togo to step #10 "Querying Data at NCBI"
Update, retract, or replace these records in NCBI databases, if necessary:

Go togo to step #11 Data curation protocol