DETERGENT CITRIC ACID TECHNIQUE FOR ISOLATING NUCLEAR AND CYTOPLASMIC FRACTIONS CONTAINING UNDEGRADED RNA FROM CELLS OF XENOPUS-LAEVIS

A simple technique is described for fractionating cells of X. laevis using Triton X-100 and citric acid. Cells were treated for 1 min with a hypotonic buffer containing 0.5% Triton X-100 and RNase inhibitors. The cells were then disrupted by adding citric acid to a final concentration of 1% for 2 min and the nuclei were pelleted by centrifugation. The detergent-citric acid fractionation technique was used to obtain pure nuclear and cytoplasmic fractions containing undegraded RNA from an epithelial cell line of X. laevis. The procedure is very gentle, eliminates RNase activity and can be performed in 5 min. The detergent-citric acid technique can easily be scaled up or down and was used successfully with mammalian and insect cell lines.