Isolation and characterization of the rRNA gene clusters of Halobacterium marismortui

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Abstract
Two rRNA operons of Halobacterium marismortui were identified and cloned into plasmid pBR322 as 10- and 20-kilobase-pair (kbp) HindIII fragments, respectively. Restriction maps of the 10-kbp clone (pHH10) and an 8-kbp HindIII-ClaI subclone (pHC8) of the other operon were established. Southern hybridization of 16S, 23S, and 5S rRNA probes to the clones demonstrated that both operons code for the three rRNA species. By S1 nuclease analysis, the transcription initiation sites, some of the processing sites within the primary transcripts, and the boundaries of the mature 16S and 23S rRNA molecules were determined. Both operons are transcribed in vivo. Comparison of the two operons indicated that they are not identical. The most striking difference between the operons is the existence of three putative transcription initiation sites in one operon (HC8) and only one such site in the other operon (HH10). The regions surrounding these 5' transcript end sites share a high level of sequence similarity to each other and to the rRNA promoter regions of other halophilic archaebacteria. Analysis of the proximal 130 nucleotides of the two 16S rRNA genes indicated greater-than-expected sequence heterogeneity. There are a 2-base-pair insertion in the HC8 16S gene and 10 additional sites of nucleotide sequence heterogeneity.