A Deoxyribonucleic Acid-associated Ribonucleic Acid from Drosophila melanogaster

Abstract

Several groups of investigators have reported the existence of natural associations between ribonucleic acid and deosyribonucleic acid (l-4).It has been postulated that the RNA of such associations is an intermediate in the translation of the genetic information encoded in the DNA (1).This report presents some observations on an RNA that is isolated in association with the DNA of Drosophila.EXPERIMENTAL PROCEDURE Materials Pancreatic RNase (five times crystallized) was obtained from California Corporation for Biochemical Research.Pancreatic DNase and snake venom phosphodiesterase were obtained from Worthington Biochemical Corporation.Aerosol OT (dioctyl sodium sulfosuccinate) was obtained from the Fisher Scientific Company.Methods Culture and Collection illethods-Awild Oregon R stock of Drosophila melanogaster which is highly inbred was used throughout these experiments.Cultures were grown on a medium containing corn meal, agar, sucrose, glucose, brewers' yeast, living yeast, propionic acid, and phosphoric acid.When the resulting progeny were to be uniformly labeled with s2P, the phosphoric acid was omitted from the medium and 5 to 10 ,nc of 32P-orthophosphate were added per ml of medium.Adult flies were collected by etherization and stored at -20".Larvae were collected from the medium by adding 60% sucrose to each bottle and decanting the larvae, which float in sucrose of this density.The larvae were washed in 10 to 12% sucrose (a density approximating that of the larvae), 30 '% sucrose (in which the larvae float), and water (in which they sink).Isolation of Nucleic Acids-The procedure was designed for 5 g of frozen flies or larvae.The procedure may be scaled up to 50 g of material but becomes cumbersome with larger amounts.All operations were carried out at 0" unless otherwise stated.Five grams of frozen flies or larvae were homogenized with a Davis-Layne Dual1 tissue grinder in 35 ml of 0.15 bf NaCl-0.015M citrate and centrifuged at 10,000 x g for 30 minutes.The 32,000 X g pellet RNA, soluble RNA, and microsomal RNA were isolated from the 10,000 x g supernatant, and the DNA with its associated RNA, from the 10,000 x g pellet.