Titration of AmPure XP Beads for Removal of Fragments <400bp
Label 11x1.5 ml tubes 40-45-50-55 etc.
to 90 (= ratio of beads to DNA) Mix 48μl NEB 100bp ladder with 1152μl Molecular Biology Water.
Pipet 100μl of mixture into each of the 11 tubes.
Vortex bottle of AmPure XP beads and dispense 800μl into fresh tube.
Using same pipettor used for step #3, pipet volume of beads into each tube (i.e., 40μl into tube labled 40, 45μl into tube labed 45, etc.)
Close lids, vortex tubes and incubate 5 minutes at room temperature.
Place tubes in MPC rack; wait 5 minutes for beads to adhere to magnet side of tubes.
Pipet off supers and discard them.
Wash the beads 2x with 500μl each 70% ethanol with 30 second incubation.
Remove all supers from tubes.
Place tubes (with caps open) into 37°C heat block to dry beads (evaporate residual ethanol).
Add 10μl Qiagen EB to each tube, close lids.
Vortex to resuspend beads.
Place tubes back into MPC rack and let beads adhere to magnet sides of tubes.
Pipet off supers (containing eluted DNA) into fresh tubes.
Run samples in 2% agarose gel (1x TAE) using 5μl DNA and 1μl 6x gel loading buffer.
Photograph results and determine what ratio of beads to DNA to use to exclude lower range of DNA.
