Design and characterization of PCR primers for detection of pathogenic Neisseriae.
Oligonucleotide polymerase chain reaction (PCR) primers directed at a group of closely related Neisserial species were designed from 16S rDNA sequences even though only a single such sequence from the targeted group was known. The amplifiable group included all Neisserial species considered pathogenic for man, including Neisseria gonorrhoea and Neisseria meningitidis. None of 43 other bacterial DNA specimens were amplified, including five non-Neisserial Neisseriaceae and three non-pathogenic Neisseriae. Another non-pathogenic Neisserial species gave a signal only at high DNA concentrations. DNA specimens from the pathogenic Neisseriae were detectable in amounts as low as 0.01 pg per PCR reaction, the approximate equivalent of a single organism, with equal sensitivity in buffer or in simple extracts of human inflammatory synovial fluids to which Neisserial DNA had been added. Simultaneously studied control specimens lacking added DNA were negative. The approach used to design these group-directed primers using only a single rDNA sequence from the targeted group by exploiting known patterns of sequence conservation among the 16S rDNA genes may prove useful for designing other similar group-directed primers. Polymerase chain reaction primers prepared in this way should prove of value in a number of areas, both investigational and clinical.
Department of Medicine, State University of New York at Stony Brook 11794-8161.
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Muralidhar
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Steinman
D001483:Base Sequence; D017931:DNA Primers; D004269:DNA, Bacterial / Q000032:analysis / Q000235:genetics*; D006801:Humans; D008969:Molecular Sequence Data; D009343:Neisseria / Q000235:genetics* / Q000302:isolation & purification; D009344:Neisseria gonorrhoeae / Q000235:genetics / Q000302:isolation & purification; D009345:Neisseria meningitidis / Q000235:genetics / Q000302:isolation & purification; D016133:Polymerase Chain Reaction; D012680:Sensitivity and Specificity; D013582:Synovial Fluid / Q000382:microbiology
D017931:DNA Primers; D004269:DNA, Bacterial