Nucleobond AX cartridge Regeneration

(1)Follow the Nucleobond purification procedure for the lysis of bacteria and the removal of the cellular debris (buffers S1, S2, and S3, and so on).

(2)Apply the supernatant to the cartridge as described, collect the entire flowthrough from the cartridge, and place the flowthrough on ice.

(3)Continue with the purification procedure, that is, add to the cartridge the recommend N3 washing buffer volume and the recommended N5 elution buffer volume and collect plasmid.

(4)Immediately flush the column with deionized water by filling it to the top rim and allowing it to completely run out. Re-equilibrate the cartridge with the prescribed amount of buffer N2, and reapply the recovered flowthrough from step (2) above. Wash and elute as usual. (NOTE 3)

(5)Repeat step (4) until the flowthrough has been depleted of plasmid. (This can be determined by checking the UV absorbance at 260 nm of the various eluates.)

(6)Combine the plasmid-containing eluates and precipitate (or precipitate the various eluates separately, if there are volume constraints).

NOTE 1: In order to avoid cross contamination, cartridge reuse is only recommended for purification of the same nucleic acid. For example, when purifying plasmids, a cartridge to be reused should be dedicated to a single plasmid.

NOTE 2: Use of this procedure may deplete the N series of buffer solutions supplied with the Nucleobond kits. Please refer to N series buffer preparation.

NOTE 3: For longer term storage, flush the column with a 50% ethanol solution after the wash with deionized water.Wrap the cartridge in plastic wrap and store at 4C until needed.