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Ultra-Long DNA Sequencing Kit
SQK-ULK001
$1,200.00

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

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 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.

The kit includes a new type of Rapid Adapter (RAP F), which is designed such that its consumption of fuel during a sequencing run is reduced significantly (over RAP), resulting in much better maintenance of enzyme speed. As such, the new adapter results in higher sequencing output.

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.

image2021-9-8 13-28-7 (1)

The Ultra-Long DNA Sequencing Kit features:

FeatureProperty
Preparation time~800 minutes
Input requirement• Tissue culture cells (e.g. GM24385): 6 million
• Peripheral blood mononuclear cells: 6 million
PCR RequiredNo
FragmentationTransposase-based
Read lengthR9.4.1 flow cells: 50–100+ kb N50, reads up to 4+ Mb
Read type produced1D
Typical throughput★★★
Associated protocolsUltra-Long DNA Sequencing Kit (SQK-ULK001)
Multiplexing optionsFlow Cell Wash Kit
Pack sizeSix reactions where each reaction supports three separate loads of either PromethION Flow Cells or 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.

Shipping and logistics:

Flow cells and kits are shipped together at 2–8°C.

Upon receipt, please place the right product in the right long-term storage location.

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 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 and sequencing.

Updated ULK protocol workflow V3 1

Protocols

Protocol for the Ultra-Long DNA Sequencing Kit (SQK-ULK001)

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

What's in the box

The Ultra-Long DNA Sequencing Kit contains sufficient reagents to generate six reactions, where each reaction supports three separate loads of either PromethION Flow Cells or MinION Flow Cells.

SQKULK001 Content in the box

NameAcronymCap colourNo. of tubesFill volume per vial (µl)
Rapid AdapterRAP FGreen140
Fragmentation MixFRABrown150
FRA Dilution BufferFDBClear11,600
Elution BufferEBBlack11,500
Sequencing BufferSQBRed11,500
Priming TetherPTPurple1600
Flush BufferFBWhite cap, light blue stripe on label37,200
L Fragment BufferLFBWhite cap, orange stripe on label26,500

3rd party materials

Consumables

  • NEB Monarch® HMW DNA Extraction Kit for Tissue (Cat # T3060)
  • 10 mM Tris.HCl, 1 mM EDTA pH 9
  • Triton X-100 0.5% v/v
  • Spermine tetrahydrochloride (Sigma-Aldrich, cat # 85605 1-G)
  • RBC Lysis Solution (QIAGEN, cat # 158904) for isolating PBMCs
  • Isopropanol
  • Ethanol
  • Qubit dsDNA BR Assay Kit (ThermoFisher, Q32850)
  • Qubit Assay tubes
  • 1.5 ml Eppendorf DNA LoBind tubes
  • 2 ml Eppendorf DNA LoBind tubes
  • 5 ml Eppendorf DNA LoBind tubes
  • 15 ml Falcon tubes

Equipment

  • Magnetic separator, suitable for 1.5 ml Eppendorf tubes
  • Thermal cycler or heat block at 75°C
  • Microfuge
  • Vortex mixer
  • P1000 pipette and wide-bore pipette tips
  • P200 pipette and wide-bore pipette tips
  • P200 pipette and tips
  • Ice bucket with ice
  • Timer

Optional Equipment

  • Qubit fluorometer (or equivalent for QC check)

Compatibility

The Ultra-Long DNA Sequencing Kit can be used together with:

Kits

  • Flow Cell Wash Kit (EXP-WSH004)

Flow cells

  • FLO-MINSP6
  • FLO-MIN106D
  • FLO-MIN111
  • FLO-PRO002

Devices

  • MinION Mk1B
  • MinION Mk1C
  • GridION
  • PromethION