PCR-DNA Probe Assays for Prevotella intermedia and Prevotella nigrescens

Christian Mouton Christian.Mouton at greb.ulaval.ca
Fri May 9 08:59:40 EST 1997


Here is the abstract of a paper to be published in Journal of Clinical
Microbiology:

PCR-DNA Probe Assays for  Identification and Detection of Prevotella
intermedia sensu stricto and Prevotella nigrescens
Emmanuelle Guillot and christian Mouton

The purpose of this study was to construct PCR-DNA probe assays specific
for Prevotella intermedia sensu stricto and Prevotella nigrescens  based on
the ability of random amplified polymorphic DNA (RAPD) fingerprinting to
generate species-specific markers. The strategy included four steps: (i)
construction of first generation DNA probes from a 850-bp RAPD marker for
P. intermedia sensu stricto and a 1,300-bp RAPD marker for P. nigrescens,
(ii) cloning and sequencing of each RAPD marker, (iii) design of primer
pairs flanking specific internal sequences of 754 bp for P. intermedia
sensu stricto and ca. 1,100 bp for P. nigrescens, and (iv) synthesis (by
PCR amplification) and digoxigenin labeling of quantities of DNA probes 754
and ca. 1,100 bp in size. The PCR-DNA probe assays combine either PCR
amplification of a 754-bp specific sequence in the genomic DNA of strains
of P. intermedia sensu stricto and hybridization with the 754-bp
digoxigenin-labeled probe or amplification of a ca. 1,100-bp sequence of P.
nigrescens and hybridization with the ca. 1,100-bp probe. Specific
hybridization was observed with the amplified DNAs from 25 strains of P.
intermedia and 24 strains of P. nigrescens, and no reaction with the PCR
products from 20 foreign species. The PCR-DNA probe assays described here
should allow a highly specific and sensitive detection of P. intermedia
sensu stricto and P. nigrescens in mixed infections.

Christian Mouton, DCD, DSO
Groupe de Recherche en Ecologie Buccale
Faculte de medecine dentaire, Universite Laval
Quebec (Quebec) G1K 7P4  CANADA
tel. (418)656-5872;  fax. (418)656-2861








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