Information
This kit is recommended for users who:
- Are working with ultra-high molecular weight DNA and would like read lengths with an N50 >50 kb
- Require an end-to-end workflow to sequence ultra-high molecular weight DNA
This is an Early Access product.
For more information about our Early Access programmes, please see this article on product release phases.
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.
The Ultra-Long DNA Sequencing Kit V14 features:
| Feature | Property |
|---|
| Preparation time | ~200 minutes plus an overnight elution |
| Input requirement | - 6 million PBMCs isolated from blood
- 6 million cells isolated from frozen tissue |
| PCR requirement | No |
| Fragmentation | Transposase-based |
| Kit chemistry | Kit 14 (V14) |
| Read length | 50–100+ kb N50, reads up to 4+ Mb |
| Associated protocols | Ultra-Long DNA Sequencing Kit V14 (SQK-ULK114) |
| Pack size | 6 reactions where each reaction supports 3 separate loads of PromethION Flow Cells and 6 MinION Flow Cells |
| Stability | Shipped at 2–8°C
Long-term storage: -20°C
Oxford Nanopore Technologies deem the useful life of the product to be 3 months from receipt by the customer. |
The Ultra-Long DNA Sequencing Kit V14 offers a means of preparing ultra-high molecular weight (uHMW) DNA for sequencing, which has shown to give N50s >50 kb and reads up to 4+ Mb. The kit is based on transposase chemistry: the transposase simultaneously cleaves template molecules and attaches tags to the cleaved ends. Rapid sequencing adapters are then added to the tagged ends. The last step is an overnight elution of the DNA library.
This kit has been updated to use our newest Kit 14 chemistry which includes improved modal raw read sequencing accuracies with higher output on our latest nanopore: R10.4.1. Our flow cell priming and sequencing reagents have been reformulated to be compatible with the Kit 14 chemistry upgrade and R10.4.1 nanopore.
Kit 14 also includes previous updates such as higher capture rate of DNA to enable lower flow cell loading amounts, and fuel fix technology, allowing users to run longer experiments without the need for fuel addition during the run.
Specific to the Ultra-Long Sequencing Kit V14, we now supply the Extraction EB (EEB) and Precipitation Buffer (PTB) prepared for customer use and updated the Flush Tether UL (FTU) for Kit 14 compatibility. A Precipitation Star (PS) is also provided in this kit to aid DNA precipitation and recovery.
For an end-to-end workflow, we have used the NEB Monarch® HMW DNA Extraction Kit for Tissue to extract uHMW gDNA, enabling users to extract and sequence their sample in just one day.
Validated inputs for the end-to-end protocol are:
- 6 million human cells isolated from culture (e.g. GM24385)
- 6 million peripheral blood mononuclear cells (PBMCs) from blood
These inputs produce high read N50s and, when paired with the Flow Cell Wash Kit (EXP-WSH004), deliver high outputs on both MinION and PromethION.
Shipping and logisitics
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 protocol outlines a complete workflow from extracting gDNA from cell culture or purified cells from whole blood using the NEB Monarch® HMW DNA Extraction Kit for Tissue to the sequencing of uHMW gDNA using the Ultra-Long DNA Sequencing Kit V14 in a day.
At the heart of the sequencing kit is a transposase which simultaneously cleaves template molecules and attaches tags to the cleaved ends. Rapid Sequencing Adapters are then added to the tagged ends, followed by an overnight elution before loading the DNA library onto a flow cell for sequencing.
What's in the box
The Ultra-Long Sequencing Kit V14 contains sufficient reagents to generate six reactions, where each reaction supports three separate loads of either PromethION or MinION and GridION flow cells.
| Name | Acronym | Cap colour | Number of vials | Fill volume per vial (µl) |
|---|
| Rapid Adapter | RA | Green | 1 | 40 |
| Fragmentation Mix | FRA | Amber | 1 | 50 |
| FRA Dilution Buffer | FDB | Clear | 1 | 1,600 |
| Elution Buffer | EB | Black | 2 | 1,500 |
| Extraction EB | EEB | Orange | 3 | 1,700 |
| Sequencing Buffer UL | SBU | Red | 2 | 1,000 |
| Loading Solution UL | LSU | White cap, pink label | 1 | 200 |
| Flush Tether UL | FTU | Purple | 1 | 600 |
| Flow Cell Flush | FCF | Clear cap, light blue label | 2 | 15,500 |
| Precipitation Buffer | PTB | Blue | 2 | 1,700 |
| Precipitation Star | PS | Yellow | 6 | 1 star |
Compatibility
The Ultra-Long DNA Sequencing Kit V14 (SQK-ULK114) can be used together with:
Kits:
- Flow Cell Wash Kit (EXP-WSH004)
- EEB Expansion (EXP-EEB001)
- Ultra-Long Auxiliary Vials (EXP-ULA001)
Flow cells:
R10.4.1:
Devices:
- MinION Mk1B
- MinION Mk1C
- GridION
- PromethION