NextSeq 550 Whole Genome Sequencing Workflow Guide
Explore the detailed sequencing workflow of SARS-CoV-2 whole genome sequencing using the Illumina NextSeq 550 system. Learn about the machine specifications, kit components, and step-by-step instructions from sample sheet creation to data analysis. Get insights into the washing, loading, cleaning, and QC metrics procedures involved in the sequencing process.
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Presentation Transcript
Module 3 SARS-CoV-2 whole genome sequencing Sequencing using NextSeq 550 Dr Arshad Ismail Zama Khumalo Thabo Mohale National Institute for Communicable Diseases, South Africa Date : 19 May 2021
Sequencing Workflow Sequencing Washing of the Machine Step 1 Create Sample Sheet Step 2 Loading of Libraries Step 3 Run - QC Metrics Step 4
NextSeq 550 Specifications Run Time Maximum output Maximum Reads per Run Maximum Read Length 12 30 hrs 120 Gb 400 million 2 150 bp
Sequencing Kit Components 1. Buffer Cartridge 1 2. Loading Cartridge 2 3 3. Flow cell
STEP 1 Washing of the Machine Quick Wash Quick wash is performed before every sequencing Run A wash solution of 40 ml of laboratory-grade water mixed with 20 l Tween 20 (0.05% Tween 20 final concentration) Wash takes about 20 minutes
STEP 2 Sample Sheet Loading a sample sheet onto the NextSeq 550
STEP 3 Cleaning & inspecting of the Flow Cell Illumina NextSeq Control Software Loading Flow cell in the compartment Loading of Buffer Cartridge
STEP 4 Loading of Libraries Loading Libraries into Reagent Cartridge Pierce foil with a new pipette on the well-marked Load Libraries Here Add libraries (650 l x2 ) to the bottom of the well Gentle tap the cartridge Slide the reagent cartridge into the reagent compartment until the cartridge stops, Close the reagent compartment door
Data analysis This concludes the wetlab portion of the Illumina CovidSeq workflow In the next session - in silico part of DNA sequencing, where softwares are used to assemble and analyze the raw data output of the next-generation sequencers
Thank you This programme is being funded by the UK Department of Health and Social Care. The views expressed do not necessarily reflect the UK Government s official policies.
