Staining of Bacteriophage Nucleic Acids with Acridine Orange

Abstract

THE use of the fluorescent stain acridine orange for the quick identification of viral nucleic acids has been described by Mayor and Hill1. In this valuable technique colour differences permit differentiation between double-stranded deoxyribonucleic acid (DNA), on one hand (fluoresces green), and ribonucleic acid (RNA) and single-stranded DNA on the other (fluoresces red). The latter nucleic acids can be separated by their respective sensitivities to RNase and DNase. While the procedure of Mayor and Hill is simple compared with the techniques of biochemical analysis, it involves numerous treatments and the use of fluorescence microscopy. Furthermore, acridine orange staining is particularly sensitive to pH levels, and, because of this, some difficulty may be experienced in obtaining the correct colours. In addition, the procedure does not differentiate between single-stranded DNA and RNA, using the stain alone. This communication describes some improvements and modifications with these points in mind. Direct viewing under ultra-violet light is substituted for the fluorescence microscope, the pre-staining treatment is simplified, and possible unreliability due to pH sensitivity is eliminated.