-
Introduction to Rapid Sequencing Kit V14 (SQK-RAD114)
This protocol describes the step-by-step instructions to complete a rapid sequencing of genomic DNA using the Rapid Sequencing Kit V14 (SQK-RAD114). This protocol uses our most recent Kit 14 chemistry and is optimised for a fast library preparation and requires minimal laboratory equipment.
It is highly recommended that a Lambda control experiment is completed first to become familiar with the technology.
Steps in the sequencing workflow:
Prepare for your experiment
You will need to:
- Extract your DNA, and check its length, quantity and purity using the Input DNA/RNA QC protocol.
The quality checks performed during the protocol are essential in ensuring experimental success. - Ensure you have your sequencing kit, the correct equipment and third-party reagents
- If not already installed, download the software for acquiring and analysing your data
- Check your flow cell(s) to ensure it has enough pores for a good sequencing run
Library preparation
You will need to:
Library preparation step Process Time Stop option Tagmentation Tagment your DNA using the Fragmentation Mix 5 minutes - Adapter attachment Attach sequencing adapters to the DNA ends 5 minutes We strongly recommend sequencing your library as soon as it is adapted. Priming and loading the flow cell Prime the flow cell and load the prepared library for sequencing 5 minutes - Sequencing and analysis
You will need to:
- Start a sequencing run using the MinKNOW software, which will collect raw data from the device and basecall reads.
- Optional: Start the EPI2ME software and select a workflow for further analysis, e.g. metagenomic analysis or drug resistance mapping.
- Extract your DNA, and check its length, quantity and purity using the Input DNA/RNA QC protocol.