Isolation of bacteriophage lambda containing yeast ribosomal RNA genes: screening by in situ RNA hybridization to plaques

Abstract

We have developed an in situ hybridization technique which can be used to screen large numbers of hybrid bacteriophage for the presence of a particular inserted DNA sequence. Plaques of hybrid phage are formed on E. coli lawns on nitrocellulose filters, and their DNA is released, denatured, and fixed directly on the filters for hybridization to radioactive RNA probes. We have used this technique to isolate a number of hybrid bacteriophage lambda which contain EcoRl restriction fragments of the ribosomal RNA genes from yeast, and have examined the DNA from several of these phage.