Information
This kit is recommended for users who:
- Want to do 16S sequencing
- Are interested in genus-level bacterial identification
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.
Barcoding or multiplexing is useful when the amount of data required per sample is less than the total amount of data that can be generated from a single flow cell: it allows a user to pool multiple samples and sequence them together, making more efficient use of the flow cell.
Multiplexing samples onto one flow cell can reduce the cost per sample for a user. Below are some worked examples:
| No barcodes | 12 barcodes | 24 barcodes | 24 barcodes (Flongle) |
|---|
| Flow cell price | $500 | $500 | $500 | $90 (Flongle flow cell) |
| Library price | $90 | $120 | $150 | $150 |
| Price per sample | $590 | $51.67 | $27.08 | $10 |
The 16S Barcoding Kit offers a method of amplifying the entire ~1,500 bp 16S rRNA gene from extracted gDNA, followed by library preparation. By narrowing down to a specific region of interest, a user can see all the organisms in the sample without sequencing unnecessary regions of the genome, making the identification quicker and more economical. On the other hand, analysing the full 16s rRNA gene is more informative than analysing a subset of exons.
The 16S Barcoding Kit features:
| Feature | Property |
|---|
| Preparation time | 10 minutes + PCR |
| Input requirement | <10 ng gDNA per sample |
| PCR Required | Yes |
| Fragmentation | Not required |
| Read length | Full-length 16S gene (~1.5 kb) |
| Kit chemistry | Kit 9 |
| Associated protocols | Rapid sequencing amplicons - 16S barcoding (SQK-16S024) |
| Multiplexing options | • 16S Barcoding Kit 1-24
• Flow Cell Wash Kit |
| Pack size | 6 reactions |
| 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 DNA is amplified by PCR using specific 16S primers (27F and 1492R) that contain 5’ tags which facilitate the ligase-free attachment of Rapid Sequencing Adapters.
The kit is supported by the EPI2ME FASTQ 16S workflow, which can be used to analyse data from the 16S protocol. Deconvolution of barcoded sequencing data is supported by Guppy and EPI2ME which classify the barcode sequence and sort reads into corresponding folders.
Shipping and logistics:
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 DNA is amplified by PCR using specific 16S primers (27F and 1492R) that contain 5’ tags which facilitate the ligase-free attachment of Rapid Sequencing Adapters. The kit contains 24x barcoded primer pairs.
The kit is supported by the EPI2ME 16S-BLAST workflow, which can be used to analyse data from the 16S protocol. Deconvolution of barcoded sequencing data is supported by Guppy and EPI2ME, which classify the barcode sequence and sort reads into corresponding folders.
Protocols
Rapid sequencing amplicons - 16S barcoding (SQK-16S024)
To access the protocol, you will need to register for a Nanopore Community account.
What's in the box
The 16S Barcoding Kit 1-24 contains 24 unique barcodes and sufficient reagents to generate six sequencing libraries.
| Name | Acronym | Cap colour | No. of vials | Fill volume per vial (μl) |
|---|
| 16S Barcode Primers 01-24 | 1-24 | - | 2 plates, 3 strips per plate | 15 μl per well |
| Rapid Adapter | RAP | Green | 1 | 10 |
| Sequencing Buffer | SQB | Red | 1 | 300 |
| Loading Beads | LB | Pink | 1 | 360 |
The Flow Cell Priming Kit Expansion Pack is also supplied.
| Name | Acronym | Cap colour | No. of vials | Fill volume per vial (μl) |
|---|
| Flush Buffer | FB | Blue | 6 | 1,170 |
| Flush Tether | FLT | Purple | 1 | 200 |
Compatibility
The PCR Sequencing Kit can be used together with:
Kits
- RAP Top-up Kit (EXP-RAP001)
- Flow Cell Wash Kit (EXP-WSH004)
Flow cells
Devices
- MinION Mk1B
- MinION Mk1C
- GridION
- Flongle