- Materials
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- Elution Buffer from the Oxford Nanopore kit (EB)
- Precipitation Buffer (PTB)
- Consumables
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- 1.5 ml Eppendorf DNA LoBind tubes
- Equipment
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- Centrifuge
- Microfuge
- Hula mixer (gentle rotator mixer)
- P200 pipette and tips
- P1000 pipette and tips
- Wide-bore pipette tips
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Thaw the kit components at room temperature, spin down briefly using a microfuge and mix by vortexing as indicated by the table below:
Reagent Thaw at room temperature Briefly spin down Mix well by pipetting Precipitation buffer (PTB) ✓ ✓ ✓ Elution Buffer (EB) ✓ ✓ ✓ Once thawed, keep all the kit components on ice.
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Using a regular pipette tip, add 500 µl of Precipitation Buffer (PTB) to the sample.
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Mix the sample by rotating on a Hula Mixer (rotator mixer) for 20 minutes at 3 rpm.
Visually inspect to check the DNA has precipitated, forming a glassy white mass.
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Centrifuge the sample at 1000 x g for 1 minute.
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Using a regular pipette tip, carefully remove the supernatant from the tube, taking care not to aspirate the DNA pellet.
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Centrifuge the sample at 1000 x g for 1 minute.
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Using a regular pipette tip, carefully remove any residual supernatant from the tube, taking care not to aspirate the DNA pellet.
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Using a regular pipette tip, add 300 µl of Elution Buffer (EB) to the tube containing the DNA. Incubate overnight at room temperature, for a minimum of 12 hours.
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Gently mix the DNA library by slowly pipetting 10 times with a wide-bore pipette tip.
Thorough but gentle resuspension of DNA is required to prevent heterogeneity in the sample.