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Rapid PCR Barcoding Kit features
This kit is recommended for users who:
- wish to multiplex samples to reduce price per sample
- have a low starting amount of DNA
- require a simple library preparation procedure
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Introduction to the Rapid PCR Barcoding protocol
This protocol describes how to carry out rapid low input barcoding of genomic DNA using the Rapid PCR Barcoding Kit (SQK-RPB004). There are 12 unique barcodes, allowing the user to pool up to 12 different samples in one sequencing experiment.
Steps in the sequencing workflow:
Prepare for your experiment
You will need to:
- Extract your DNA, and check its length, quantity and purity.
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
- Download the software for acquiring and analysing your data
- Check your flow cell to ensure it has enough pores for a good sequencing runLibrary preparation
You will need to:
- Tagment your DNA using the Fragmentation Mix in the kit
- PCR using the barcoded primer supplied in the kit
- Attach sequencing adapters supplied in the kit to the DNA ends
- Prime the flow cell, and load your DNA library into the flow cellNote that after the PCR, the average length of DNA fragments will be 2-3 kb.
Sequencing and analysis
You will need to:
- Start a sequencing run using the MinKNOW software, which will collect raw data from the device and convert it into basecalled reads
- Start the EPI2ME software and select the barcoding workflow