A restriction map of the T4 transfer RNA gene cluster

Abstract

Bacteriophage T4 cytosine-containing DNA obtained from the T4 mutant (dCMP hydroxymethylase −, dCTPase −, endonuclease IV −) was cleaved by restriction endonucleases. Using two independent methods, a restriction map of the T4 tRNA gene cluster region was constructed. In situ hybridization of the DNA fragments with each of the 32P-labeled T4 tRNAs ordered and positioned the tRNA genes. There are two subclusters of T4 tRNA genes in the tRNA cluster region (Wilson et al., 1972), which are separated by a distance of about 600 base-pairs. The order of genes with respect to transcription is tRNA Gln, tRNA Leu, tRNA Gly, tRNA Pro, tRNA Ser, tRNA Thr, tRNA Ile in one subcluster, and tRNA Arg, species II RNA and species I RNA for the other subcluster. The overall size of the T4 tRNA gene cluster is about 1600 base-pairs. To further establish the restriction endonuclease map of this region the 5000 base-pair EcoRI fragment, which contains one of the tRNA subclusters, was cloned into a bacteriophage lambda vector (see the following paper). Using the cloned fragment a map for the cleavage sites of 14 restriction endonucleases was constructed by direct methods. The maps obtained by the two procedures are in good agreement.