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Nanopore sequencing offers advantages in all areas of research. Our offering includes DNA sequencing, as well as RNA and gene expression analysis and future technology for analysing proteins.

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Oxford Nanopore TechnologiesFully scalable, real-time DNA/RNA sequencing technology
Oxford Nanopore DiagnosticsLamPORE – rapid, low-cost, scalable detection of SARS-CoV-2
PCR-cDNA Sequencing Kit
SQK-PCS109

Identification and quantification of full-length transcripts with highest throughput

 Includes a Flow Cell Priming Kit ll
£485.00

Information

This kit is highly recommended for users who:

  • have a limited amount of input material
  • want to optimise their sequencing experiment for throughput
  • would like to identify and quantify full-length transcripts
  • are interested in differential gene expression
  • want to characterise and quantify isoforms, splice variants and fusion transcripts

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 PCR-cDNA Sequencing Kit is used to prepare cDNA for nanopore sequencing from an input of as low as 1 ng poly-A+ RNA. Users who do not have poly-A+ enriched RNA can use 50 ng of total RNA but additional optimisation may be required.

Taking full-length poly-A+ RNA, complementary strand synthesis and strand switching are performed using kit-supplied oligonucleotides. dscDNA is then generated by PCR amplification using primers that contain 5’ tags which facilitate the ligase-free attachment of Rapid Sequencing Adapters.

The PCR-cDNA Sequencing Kit features:

FeatureProperty
Preparation time165 minutes
165 minutes1 ng poly-A+ RNA (or 50 ng total RNA)
Reverse transcription requiredYes
PCR requiredYes
Read lengthEnriched for full-length cDNA during PCR
Read type produced1D
Typical throughput★★★
Typical number of reads7–12+ million full-length mapped readsper flow cell on MinION/GridION; 50+ million mapped reads per flow cell on PromethION
Associated protocolscDNA-PCR sequencing

Sequence-specific cDNA-PCR sequencing
Multiplexing options• PCR-cDNA Barcoding Kit
• Flow Cell Wash Kit
Pack size6 reactions
StabilityShipped 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.

Further considerations

The PCR-cDNA Barcoding Kit (SQK-PCB109) can be used instead of the PCR-cDNA Sequencing Kit to multiplex up to 12 different RNA samples into a single flow cell.

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

Taking full-length PolyA+ RNA, complementary strand synthesis and strand switching are performed using kit-supplied oligonucleotides. dscDNA is then generated by PCR amplification using primers that contain 5’ tags which facilitate the ligase-free attachment of Rapid Sequencing Adapters.

2019 01 07 SQK PCS109 workflow v1

Protocols

cDNA-PCR sequencing

Sequence-specific cDNA-PCR sequencing

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

What's in the box

The PCR-cDNA Sequencing Kit contains sufficient reagents to generate six sequencing libraries.

2019 01 25 SQK-PCS109 contents v2

NameAcronymCap colourNo. of vialsFill volume per vial (μl)
VN PrimerVNPBlue1200
Strand Switching PrimerSSPPink1160
cDNA PrimercPRMGrey140
Elution BufferEBBlack2200
Rapid AdapterRAPGreen110
Sequencing BufferSQBRed2300
Sequencing TetherSQTPurple110
Loading BeadsLBPink1360

The Flow Cell Priming Kit is also supplied.

FLPEXP-FLP002({{media url= "EXP-FLP002")

NameAcronymCap colourNo. of vialsFill volume per vial (μl)
Flush BufferFBBlue61,170
Flush TetherFLTPurple1200

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)
  • Freshly prepared 70% ethanol in nuclease-free water
  • Agencourt AMPure XP beads
  • 10 mM dNTP solution (e.g. NEB cat # N0447)
  • LongAmp Taq 2X Master Mix (e.g. NEB cat # M0287)
  • Maxima H Minus Reverse Transcriptase (200 U/µl) with 5x RT Buffer (ThermoFisher, cat # EP0751))
  • RNaseOUT™, 40 U/μl (Life Technologies, cat # 10777019)
  • Exonuclease I (NEB, cat # M0293)

Equipment

  • Hula mixer (gentle rotator mixer)
  • Magnetic rack
  • Microfuge
  • Vortex mixer
  • Ice bucket with ice
  • Timer
  • Thermal cycler
  • P1000 pipette and tips
  • P100 pipette and tips
  • P20 pipette and tips
  • P10 pipette and tips
  • P2 pipette and tips
  • Pre-chilled freezer block at -20° C for 200 µl tubes (e.g. Eppendorf cat # 022510509)

Optional equipment

  • Agilent Bioanalyzer (or equivalent)
  • Qubit fluorometer (or equivalent for QC check)
  • Eppendorf 5424 centrifuge (or equivalent)

Compatibility

The PCR-cDNA Sequencing Kit can be used together with:

Kits

  • Flow Cell Wash Kit (EXP-WSH004)

Flow cells

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

Devices

  • MinION Mk1B
  • MinION Mk1C
  • GridION
  • PromethION