Information

This kit is recommended for users who:

• wish to multiplex samples to reduce price per sample
• need a PCR-free method of multiplexing to preserve additional information such as base modifications
• require a short preparation time
• have limited access to laboratory equipment

Please note that to use this kit, you will need to purchase additional 3rd party reagents: see the "3rd Party Materials" tab for more detail.

Barcoding or multiplexing is useful when the amount of data required per sample is less than the total amount of data that can be generated from a single flow cell: it allows a user to pool multiple samples and sequence them together, making more efficient use of the flow cell.

Multiplexing samples onto one flow cell can reduce the cost per sample for a user. Below are some worked examples:

The Rapid Barcoding Kit generates barcoded sequencing libraries from extracted gDNA in 10 minutes using a simple 2-step protocol. At the heart of the kit is a transposase which simultaneously cleaves template molecules and attaches barcoded tags to the cleaved ends: there are 12 unique barcoded tags in the kit. Barcoded samples are pooled and Rapid Sequencing Adapters are then added to the tagged ends.

The Rapid Barcoding Kit features:

The kit is optimised for simplicity and speed, rather than for obtaining maximum throughput. Due to the simple nature of the workflow and the fact that little sample manipulation is required (e.g. minimal pipetting steps and no clean-ups), some very long reads can be achieved with this kit, despite the required transposase fragmentation. However, in order for long reads to be observed in sequencing, long fragments need to be present in the sample in the first place.

Typical read length histogram when preparing an E.coli library with the Rapid Barcoding Kit.

The workflow is PCR-free and therefore removes PCR bias and retains base modification information which can be analysed using tools developed in the Nanopore Community.

Deconvolution of barcoded sequencing data is supported by the MinKNOW and Guppy software packages, which classify the barcode sequence and sort reads into corresponding folders.

Further considerations:

The Rapid Barcoding Kit recommends a total input of 400 ng gDNA, and is optimised for samples which contain long fragments (>30 kb). Addition of less than 400 ng, or shorter fragments could compromise sequencing throughput and read length. For samples where only lower inputs are possible, the Low Input by PCR Barcoding Kit and Rapid Low Input by PCR Barcoding Kit are available.

Additional Rapid Sequencing Adapter (RAP) can be purchased (EXP-RAP001) to support the use of the Rapid Barcoding Kit.

Shipping and logistics:

Flow cells and kits are shipped together in an environmentally friendly temperature-controlled shipping box.

Products are shipped to customers within the USA and EU Monday to Thursday. Shipments to Canada, Norway, Korea and Japan are expedited Monday to Wednesday; with Australia and New Zealand leaving our warehouses on a Friday. Shipments to the rest of the world are made on Mondays to allow the full working week for packages to arrive.

The delivery charges are calculated when a quote is raised or during checkout. Once an order is made, the delivery ID and delivery information can be tracked in the Store.

Workflow

The Rapid Barcoding Kit generates barcoded sequencing libraries from extracted gDNA in 10 minutes using a simple 2-step protocol. At the heart of the kit is a transposase which simultaneously cleaves template molecules and attaches barcoded tags to the cleaved ends: there are 12 unique barcoded tags in the kit. Barcoded samples are pooled and Rapid Sequencing Adapters are then added to the tagged ends.

Protocols

Rapid Barcoding Sequencing (SQK-RBK004)

To access the protocol, you will need to register for a Nanopore Community account.

Safety and legal

Safety Data Sheet for Rapid Barcoding Kit (SQK-RBK004)

Safety Data Sheet for Flow Cell Priming Kit

What's in the box

The Rapid Barcoding Kit contains twelve unique barcodes and sufficient reagents to generate six sequencing libraries.

The Flow Cell Priming Kit is also supplied.

3rd party materials

Consumables

• 1.5 ml Eppendorf DNA LoBind tubes
• 0.2 ml thin-walled PCR tubes
• Nuclease-free water (e.g. ThermoFisher, cat # AM9937)

Optional consumables

• Agencourt AMPure XP beads
• Freshly-prepared 70% ethanol in nuclease-free water
• 10 mM Tris-HCl pH 8.0 with 50 mM NaCl