• Place tubes on a magnet holder for 2 min (or until the liquid is clear).
• Transfer 50μl in new tube for second clean-up.
6. Clean up with AMPure beads
• 50μl after ligation.
• + 55μl AMPure XP magnetic beads.
• Gently pipet up and down 10Â.
• 5 min at room temperature.
• Place tubes on a magnet holder for 2 min (or until the liquid is clear).
• Discard supernatant.
• 2 Â 500μl of 70% ethanol for 30 s, wash magnetic beads, discard supernatant.
• Air dry the beads on magnetic stand at RT for 2–5 min or at 37
C in a heatblock.
• Elute with 20 μl H2O, pipette the entire volume up and down 10 times to mix
thoroughly.
• 2 min at room temperature.
• ↺ 13,000 rpm, 1 s.
• Place tubes on a magnet holder for 2 min (or until the liquid is clear).
• Transfer 20μl in new tube for PCR (0.5 ml PCR tube).
7. Enrich DNA Fragments
Master mix final PCR (ChIP)
1 x [μl]
x [μl]
PCR Primer Cocktail[7]
5.00
PCR Primer Master Mix[8]
25.00
30.00
20μl DNA sample
+30μl master mix
50μl.
PCR-cycler:
98
C
3 0 s
4Â
98
C
1 0 s
60
C
3 0 s
72
C
3 0 s
72
C
5 min
4
C
1
Appendix
203