Use of the 16S-23S ribosomal genes spacer region in studies of prokaryoticdiversity

The description of microbial diversity by molecular culture-independent tec hniques most often involves the amplification of the 16S rRNA by PCR gene a nd either analysis of the diversity of amplified molecules (community finge rprinting) that allows the simultaneous study of many samples or the clonin g and sequencing of a significant amount of amplification products. The fac t that between the 16S and the 23S genes in the ribosomal operon there is a spacer extremely Variable in both sequence and length provides an excellen t tool to simplify both approaches. The spacer can be amplified almost as e asily as the 16S rDNA taking advantage of conserved nucleotide stretches at the 5' end of the 23S gene and the amplicon can contain different amounts of the 16S rDN4 choosing primers at the different conserved areas within th is gene. Identified by the acronym RISA (rDNA internal spacer analysis), th e spacer addition provides a marker of highly variable size allowing standa rd separation of the amplification products and the sequence of this hyperv ariable region is useful in the fine discrimination of operational taxonomi c units. (C) 1999 Elsevier Science B.V. All rights reserved.