Information

The Direct RNA Sequencing Kit (SQK-RNA004) is highly recommended for users who:

• Would like to remove PCR bias
• Are exploring attributes of native RNA such as modified bases

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.

Also included are the MinION/GridION RNA Flow Cells (FLO-MIN004RA). This is our RNA-specific flow cell and paired with the Direct RNA Sequencing Kit (SQK-RNA004), improved sequencing accuracy and output is generated for RNA libraries.

This is an Early Access product.
For more information about our Early Access programmes, please see this article on product release phases.

The Direct RNA Sequencing Kit features:

The Direct RNA Sequencing Kit (SQK-RNA004) is used to prepare and sequence native RNA without conversion to cDNA. Inputs include poly(A)-tailed RNA or total RNA, such as eukaryotic mRNA and viral RNA.

This kit upgrade includes increased sequencing output and improved modal raw read accuracy on our new RNA flow cell (FLO-MIN004RA and FLO-PRO004RA). The flow cell priming reagents are also included as part of the sequencing kit and have been reformulated to be compatible on the new RNA flow cell. Other updates include fuel fix technology, allowing users to run longer experiments without the need for fuel addition during the run.

Further considerations:

In applications where Direct RNA sequencing’s key advantages (detection of modified bases and reduced bias) are not important, users may consider one of Oxford Nanopore’s cDNA kits which currently provide the highest output per run.

Shipping and logistics:

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

Flow cells and kits are shipped together at 2–8°C. The packaging is designed to protect the flow cells from freezing. Flow cells are stable at ambient temperatures and stored refrigerated for longer periods. Sequencing kits are stable at room temperature for up to seven days, so users should not be concerned about their products should they arrive at ambient temperature.

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.

Storage and stability:

Please store your flow cells as follows:

• Room temperature: during Early Access, we ask users to store their flow cells at 2-8°C. Flow cells are stable for a few days at room temperature, however we will formalise guidance during the course of product roll-out.
• 2-8°C: flow cells can be stored, unopened, at 2-8°C for 4 weeks.

We recommend you check your flow cell before preparing your RNA library as part of the warranty process.

Flow cell returns:

The flow cell pouch provided serves as the flow cells returns packaging. Flow cells contain finite materials which, when returned to Oxford Nanopore, are responsibly recycled.

More information on how to carry out a return can be found on the Nanopore Community.

Workflow

The Direct RNA Sequencing Kit (SQK-RNA004) is used to prepare either poly(A)-tailed RNA or total RNA for sequencing by performing reverse transcription to synthesise a secondary complementary cDNA strand for stability to improve sequencing output. Sequencing adapters are then attached to the RNA-cDNA hybrid for sequencing.

Protocols

• Direct RNA sequencing (SQK-RNA004)
• Direct RNA sequencing - control experiment (SQK-RNA004)
• Direct RNA sequencing - sequence-specific (SQK-RNA004)

Safety and legal

Safety Data Sheet for Direct RNA Sequencing Kit (SQK-RNA004)

Safety Data Sheet for MinION Flow Cell RNA (FLO-MIN004RA)

Customers are required to perform a flow cell check within 4 weeks of purchase. Oxford Nanopore Technologies will replace any flow cell with fewer than 800 nanopores, when the result is reported within two days of performing the flow cell check and when the storage recommendations have been followed. The Early Access warranty on this product is 4 weeks from receipt by the customer.

There is a 9-month contract length available for all consumable purchases. For more information, please see the Nanopore Product Terms and Conditions.

What's in the box

This promo pack contains:

• 10 x MinION/GridION RNA Flow Cells (FLO-MIN004RA)
• 2 x Direct RNA Sequencing Kit (SQK-RNA004)

3rd party materials

Consumables:

• 1.5 ml Eppendorf DNA LoBind tubes
• 0.2 ml thin-walled PCR tubes
• Nuclease-free water (e.g. ThermoFisher, AM9937)
• Freshly prepared 70% ethanol in nuclease-free water
• SuperScript™ III Reverse Transcriptase (Thermo Fisher Scientific, 18080044)
• 10 mM dNTP solution (e.g. NEB N0447)
• NEBNext® Quick Ligation Reaction Buffer (NEB, B6058)
• T4 DNA Ligase 2M U/ml (NEB, M0202T/M)
• RNaseOUT™, 40 U/μl (Life Technologies, 10777019)
• Agencourt RNAClean XP beads (Beckman Coulter™, A63987)
• Qubit RNA HS Assay Kit (ThermoFisher Q32852)
• Qubit dsDNA HS Assay Kit (ThermoFisher, cat # Q32851)
• Qubit™ Assay Tubes (Invitrogen, Q32856)

Equipment:

• Hula mixer (gentle rotator mixer)
• Magnetic separator, suitable for 1.5 ml Eppendorf tubes
• Microfuge
• Vortex mixer
• Ice bucket with ice
• Timer
• Thermal cycler
• Qubit fluorometer (or equivalent for QC check)
• P1000 pipette and tips
• P200 pipette and tips
• P100 pipette and tips
• P20 pipette and tips
• P10 pipette and tips
• P2 pipette and tips
• Eppendorf 5424 centrifuge (or equivalent)

Note: The above are 3rd party reagents required for the standard prep. For other workflows using this kit, extra 3rd party reagents may be required.