Incorporation of Tritium of ³H-Arginine into DNA as the Explanation of “Late Synthesis of Protein” on the Human X Chromosome

Abstract

CHERNICK'S¹ demonstration of apparent late synthesis of protein on the X chromosome of female lymphocytes labelled with ³H-arginine¹ interested me—for several years ago I carried out a similar experiment. The studies were done on an individual with an iso-X chromosome so that there was no question of the identification of the late replicating X chromosome. Lymphocytes were grown in culture with phytohaemagglutinin and, after 72 h, 25 µCi/ml. of ³H-arginine (Schwarz BioResearch, Inc.) and 0.1 µg/ml. of colchicine were added to the culture. Three h later the cultures were collected, treated with hypotonic, fixed in acetic acid–methanol and flame-dried. They were then stained with aceto-orcein, covered with Kodak AR-10 stripping film, and autoradiographed for 48 days. The iso-X chromosome was late labelled (Fig. 1) in a pattern identical to that seen with ³H-thymidine. Similar studies with normal female lymphocytes also showed a labelling pattern that mimicked that of ³H-thymidine.