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Unlike traditional PCR, Countable PCR enables DNA recovery from the Countable Matrix for use in downstream applications such as sequencing or reamplification. This provides maximum value from each sample—saving time, resources, and precious materials. After thermal cycling, 1-butanol is added to free the amplified DNA from the compartments. The DNA is then purified using SPRIselect beads. See below for protocol details and refer to the SPRIselect manual for additional details.

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How to recover DNA from the Countable Matrix

Materials

1-Butanol (Millipore Sigma, Cat #: 19422) SPRIselect Reagent (Beckman Coulter, Cat #: B23317, B23318, B23319)

Protocol

  1. After PCR, transfer Matrix Tube Strips to a post-amplification room to minimize contamination.
  2. Gently uncap the Matrix Tube Strips taking care to avoid splatter.
  3. Using a pipette, remove the Matrix Reagents without disturbing the Countable Matrix.
  4. Add 75 µL of 1-butanol to each sample and pipette to mix. Transfer the solution to a new tube.
  5. Vortex SPRIselect beads for 30 seconds.
  6. Add SPRIselect beads. Refer to the bead-to-reaction volume ratio for the specific DNA size needed for your application. Vortex to mix.
  7. Incubate at room temperature for 5 minutes.
  8. Place the tube on a magnetic rack. Once the solution is clear, discard the supernatant.
  9. Add 200 µL of 80% ethanol. Discard the supernatant. Repeat this wash twice. Remove all residual ethanol on the final wash.
  10. Dry the SPRIselect beads at room temperature for approximately 2 minutes. Avoid over-drying, which can reduce DNA elution efficiency.
  11. Remove the tube from the magnetic rack, and resuspend the beads in 50 µL of nuclease-free water or your preferred elution buffer.
  12. Incubate at room temperature for 1 minute before placing the tubes on the magnetic rack. Once the solution is clear, transfer the supernatant to a new tube.
  13. Quantify the DNA product and continue with your downstream application.

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