Preparation of Sonicated Salmon Sperm DNA

Preparation of Sonicated Salmon Sperm DNA

Use Pharmacia #27-4564-01. With clean flamed scissors and forceps, weigh 0.25 g/50 ml conical and add 50 ml/conical of 0.02 M Tris, pH 7.6. Allow to dissolve over several days at 4°C.

Vortex. Draw up into a 20 or 30 ml syringe. Add a 25 guage needle and shear DNA by squirting through needle into fresh 50 ml conicals at 15 ml/conical.

Add 0.9 ml/conical of 5 N NaCl.

Boil in water bath (95 - 100°C) for 20 minutes.

Cool in ice water.

Vortex and then add 1 drop (from a standard transfer pipet) of 1.2 N HCl (concentrated HCl diluted 1/10) to neutralize.

Add two volumes of 100% ethanol, let stand for 20 minutes on ice.

Spin for 10 minutes in the Beckman GS-6R at 2500 rpm (4°C).

Pour off the supernatant, let dry for 5 min

Add 15 ml/tube of 0.02 M Tris, pH 7.6. Vortex.

Allow to dissolve for several days at 4°C.

Determine OD260 of a diluted aliquot to check concentration (1 OD260 = 50 µg DNA/ml), then store at -20°C.

This method is based, with permission, on an original protocol available here.

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