Related base sequences in the DNA of simple and complex organisms. II. The interpretation of DNA-RNA hybridization studies with mammalian nucleic acids.

Abstract

A comparison has been made of the properties of hybridization reactions using mouse RNA and B. subtilisRNA. By virtue of the high rates of reaction of mammalian RNA and the imperfect nature of the base pairing in the hybrids formed, it is concluded that these reactions are not locus-specific. The same cross-reaction is obtained between polynucleotides representing similar but not identical genes as is observed in the formation of mammalian DNA/DNA duplexes. Thus, saturation experiments are not accurate measurements of the fraction of the DNA responsible for the synthesis of a given mixture of RNA molecules. In fact, very different values are obtained when the incubation temperature is varied since the extent of cross-reaction of RNA with the DNA of other than parental cistrons depends on the incubation temperature. Likewise, competition experiments devoted to comparisons of different RNA preparations provide estimates of similarity which are completely dependent upon the ionic strength and the temperature of incubation. A high degree of discrimination may be attained through the use of carefully chosen reaction conditions. For example, it appears that the majority of hybridized RNA molecules from kidney and liver are distinguishable. Agreement is obtained between experiments employing simultaneous competition of two groups of RNA molecules and those in which the DNA is presaturated with unlabeled RNA before addition of the labeled RNA. In the light of these differences between hybridization reactions by RNA of simple and complex organisms, consideration is made of the applicability of the technique to the analysis of differential gene transcription in higher organisms.