The transcriptional mapping method developed previously has been used to investigate the functional organization of the ribosomal ribonucleic acid (rRNA) and transfer ribonucleic acid (tRNA) cistrons in the Bacillus subtilis genome. The sensitivities of the syntheses of these RNA species to actinomycin D indicated that (i) large polycistronic groupings of the tRNA cistrons exist, but (ii) the 16S and 23S rRNA cistrons are probably transcribed as individual units, and (iii) each 5S rRNA cistron is part of an operon containing a 23S rRNA cistron.