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Comparative Study
. 2002 Aug 1;30(15):3481-9.
doi: 10.1093/nar/gkf450.

PRIMROSE: a computer program for generating and estimating the phylogenetic range of 16S rRNA oligonucleotide probes and primers in conjunction with the RDP-II database

Kevin E Ashelford  1 , Andrew J Weightman, John C Fry
Affiliations
Comparative Study

PRIMROSE: a computer program for generating and estimating the phylogenetic range of 16S rRNA oligonucleotide probes and primers in conjunction with the RDP-II database

Kevin E Ashelford et al. Nucleic Acids Res. .

Abstract

We describe PRIMROSE, a computer program for identifying 16S rRNA probes and PCR primers for use as phylogenetic and ecological tools in the identification and enumeration of bacteria. PRIMROSE is designed to use data from the Ribosomal Database Project (RDP) to find potentially useful oligonucleotides with up to two degenerate positions. The taxonomic range of these, and other existing oligonucleotides, can then be explored, allowing for the rapid identification of suitable oligonucleotides. PRIMROSE includes features to allow user-defined sequence databases to be used. An in silico trial of the program using the RDP database identified oligonucleotides that described their target taxa with a degree of accuracy far greater than that of equivalent currently used oligonucleotides. We identify oligonucleotides for subdivisions of the Proteobacteria and for the Cytophaga-Flexibacter-Bacteroides (CFB) division. These oligonucleotides describe up to 94.7% of their target taxon with fewer than 50 non-target hits, and the authors recommend that they be investigated further. A comparison with PROBE DESIGN within the ARB software package shows that PRIMROSE is capable of identifying oligonucleotides with a higher specificity. PRIMROSE has an intuitive graphical user interface and runs on the Microsoft Windows 95/NT/2000 operating systems. It is open source and is freely available from the authors.

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Figures

Figure 1
Figure 1
Flow chart summarising the procedure followed by PRIMROSE to identify oligonucleotides. SSU_Prok.gb is a GenBank formatted file produced by the RDP containing all currently aligned 16S rRNA sequence records (currently totalling 16 277 records in release 8.1). SSU_Prok.alpha, SSU_Prok.phylo and SSU_Prok.phylo.stats are associated files, also from RDP, that supply additional phylogenetic-based information on these records. All files are available from the RDP web site at http://rdp.cme.msu.edu/html/download.html.
Figure 2
Figure 2
Using sequence data selected from the RDP, PRIMROSE has identified a range of possible oligonucleotides for the CFB division. Each listed oligonucleotide is a hypertext link to further information on the phylogenetic range of that sequence.
Figure 3
Figure 3
Further information on the degenerate oligonucleotide WCCCTTTAAACCCART (probe CFB560) which has been selected for further investigation by clicking the hypertext link shown in Figure 2. Displayed within this window is part of a phylogenetic breakdown of those RDP records targeted by this oligonucleotide.
Figure 4
Figure 4
Additional information on the oligonucleotide of interest. In this example, the program shows the range and position of the four non-degenerate versions of the degenerate probe CFB560 (12).
See this image and copyright information in PMC

References

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