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  • Cleanup of Adaptor Ligated DNA (E6285)

    Protocols

    1. Add 72 µl (1.8 x volume) of AMPure XP Beads to the sample and mix by pipetting up and down.

    2. Incubate for 5 minutes at room temperature.

    3. Pulse-spin the tube and place in a magnetic rack for approximately 2-3 minutes until the beads have collected to the wall of the tube and the solution is clear.

    4. Carefully remove and discard the supernatant without disturbing the beads that contain the DNA target.

    5. Keep the tube on the magnet and add 200 µl freshly prepared 80% ethanol. Incubate to room temp for 30 seconds and carefully remove and discard the supernatant.

    6. Repeat step 5.

    7. Keeping the tube in the magnetic rack, with the cap open, air dry the beads for 5 minutes at room temperature.
      Caution: Do not overdry the beads. This may result in lower recovery of DNA target.

    8. Remove the tube from the magnet. Resuspend the beads in 25 μl of 0.1X TE (volume may be adjusted for specific size selection protocol). Incubate for 2 minutes at room temperature.

    9. Pulse spin the tube, and place in the magnetic rack until the beads have collected to the side of the tube and the solution is clear.

    10. Transfer approximately 20 μl of the supernatant to a clean tube. Be careful not to transfer any beads.

    11. For E-Gel or Agarose gel size selection, select adaptor ligated DNA in the 190–230 bp range for 100 bp libraries and 290–330 bp range for 200 bp read lengths.

    12. Proceed to PCR Amplification of Adaptor Ligated DNA.