Express delivery and free returns within 30 days

Ultra-Long DNA Sequencing Kit V14

An overnight method to prepare ultra-high molecular weight DNA for sequencing ultra-long reads (N50 >50 kb).

This is an Early Access product

This uses our latest Kit 14 chemistry

SKU SQK-ULK114 Categorías ,
Share it :

Descripción

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 milling 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.

ULK114 workflow V1-3

Protocols

Ultra-Long DNA Sequencing Kit V14 (SQK-ULK114)

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

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.

SQK-ULK114 tubes v1

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

3rd party materials

Consumables

  • 1.5 ml Eppendorf DNA LoBind tubes
  • 2 ml Eppendorf DNA LoBind tubes
  • 5 ml Eppendorf DNA LoBind tubes
  • 15 ml Falcon tubes
  • Isopropanol, 100%
  • Ethanol, 100%
  • Phosphate Buffered Saline (PBS)
  • Qubit dsDNA BR Assay Kit (ThermoFisher, Q32850)
  • Qubit™ Assay Tubes (ThermoFisher, Q32856)
  • (Optional) Bovine Serum Albumin (BSA) (50 mg/ml) (e.g Invitrogen™ UltraPure™ BSA 50 mg/ml, cat# AM2616)

Equipment

  • Thermal cycler or heat block
  • Thermomixer
  • Hula mixer
  • Vortex mixer
  • Microfuge
  • Wide-bore pipette tips
  • P1000 pipette and tips
  • P200 pipette and tips
  • P20 pipette and tips
  • Qubit fluorometer (or equivalent)
  • Ice bucket with ice
  • Timer

Compatibility

The Ultra-Long DNA Sequencing Kit V14 (SQK-ULK114) can be used together with:

Kits:

  • Flow Cell Wash Kit (EXP-WSH004)

Flow cells:

R10.4.1:

  • FLO-MIN114
  • FLO-PRO114M

Devices:

  • MinION Mk1B
  • MinION Mk1C
  • GridION
  • PromethION

Valoraciones

No hay valoraciones aún.

Sé el primero en valorar “Ultra-Long DNA Sequencing Kit V14”

Tu dirección de correo electrónico no será publicada. Los campos obligatorios están marcados con *