Technique simple de préparation des acides désoxyribonucleiques de foie de rat

Abstract

The frozen tissue was sliced and then homogenized at 20 °C in LiCl, 2 m; lauryltrimethyl ammonium chloride (K & K No. 4484), 5 %; pronase, B grade (Calbiochem), 1 mg/ml and Tris-HCl 10 mM pH 7.5. The homogenate was left to stand 15 min at 20 °C with occasional shaking. After centrifugation at 35 000 × g for 30 min at 0 °C, the supernatant containing the crude DN A was purified by filtration on Ultrogel A 2 (LKB, Sweden). The Ultrogel A 2 column (2.5 × 45 cm) was equilibrated with a solution containing NaCl 2 m, EDTA 2.5 mM, Tris-HCl 10 mM pH 7.5. The flow rate was 3 ml cm⁻² h⁻¹. Five ml of the supernatant were placed on the column. The first peak contained highly polymerized (as demonstrated by ultracentrifugation) pure DNA (A₂₆₀/A₂₃₀ = 3.19; A₂₆₀/A₂₈₀ = 1.82). The yield was 2.26 mg of DNA/g of fresh liver.