Molecular differentiation of bacteria by PCR amplification of the 16S–23S rRNA spacer

In general, operons for prokaryotic rRNA genes contain a transcribed spacer between 16S and 23S rRNA genes. The length and sequence of this spacer are expected to be highly variable. Polymerase chain reaction (PCR) permits the direct amplification of spacer DNA from small amounts of genomic DNA. Data for a wide range of microorganisms were accumulated, and the spacer lengths varied between 280 and 1300 bp. A further differentiation was achieved by a high-resolution gel electrophoresis method, single-strand conformation polymorphism (SSCP). It was possible to differentiate 15 Mycoplasma species, to analyze mixed samples and to distinguish Streptococcus strains and Clostridium botulinum serotypes. Spacer analysis is a promising method for the differentiation of diverse and closely related species, and also useful in analysis of mixed samples.