Expression of the chromosome-coded rRNA genes of Proteus vulgaris in Escherichia coli.

Abstract

To trace the fate of heterologous rRNA operons in vivo, a complete rrn operon from P. vulgaris was transferred into the chromosome of E, coli. This was done by phage ? gt 11 and by plasmid pOM40 which promotes the integration of the cloned insert into the maiP locus. As derived from oligonucleotide analysis of 16S rRNA isolated from ribosomal 30S subunits the amount of heterologous 16S rRNA in the ribosomes of corresponding clones was determined to be about 5 % of the homologous E. coil 16S rRNA.Recently we reported the construction of recombinant plasmids by cloning ribosomal RNA operons from Proteus vulgaris in plasmid pBR322. Transformation of these plasmids in Escherichia coli lead to the formation of hybrid ribosomes in the corresponding clones (10) to which P, vulgaris 16S rRNA contributed about 25%. As demonstrated by in vitro transcription experiments (9) heterologous promoters are as efficiently recognized as the homologous promoters. In addition, in vitro translation by hybrid E, coil ribosomes, using the MS 2 system, has been shown to be as efficient as a homogeneous ribosome population from E. coil (unpublished). The aim of this study was to investigate the possibility of integrating a heterologous rRNA operon into the genome of E. coil to answer questions in the long run about (i) the maintenance of the almost perfect uniformity of the primary structure of rDNA genes located on different operons, and (ii) the fate of a heterologous ribosomal gene. The latter question is of crucial interest for modern rRNA based phylogeny (14).

Phylogenetic evidence for the relationship between the genera Mobiluncus and Actinomyces

“…Underlined re~ons mark putative target sites for Mobi~nc~-specific ofigonucleotide probes. Table 1 Estimated A, Actmomyces [12]; Pr, Propionibacterium [11]; B, Bifidobacterium (Hensieck and Stackebrandt, unpublished); S, Saccharomonospora [3]; P, Pseudonocardia [3]; C, Corynebacterium [15];…”

“…2) which shows Mobiluncus and Actinomyces to possess a common ancestry. The topography of the tree (omitting Mobiluncus) is very similar to the one derived from distance matrix analysis of a slightly different 16S rRNA data base [11,12].…”

Phylogenetic evidence for the classification of Acidothermus cellulolyticus into the subphylum of actinomycetes

“…Extraction of genomic DNA from lyophilized cells, amplification of 16S rDNA and purification of PCR products were performed as previously described [7]. The double-stranded PCR products were sequenced using primers as described previously [8] and the automated method as described by Rainey and Stackebrandt [9]. The sequence, which has been deposited under accession number X70635, was aligned against the small subunit rRNA collection of the NSF Ribosomal Database project [10] supplemented with recent releases from EMBL.…”

Propionigenium modestum: a separate line of descent within the eubacteria

“…Isolation of crude ribosomal RNA was done as published [6]. Genomic DNA was purified following Marmur's method [7] with the modifications described by Meyer and Schleifer [8].…”