Polymerase chain reaction of bacterial genomes with single universal primer: application to distinguishing mycobacteria species.
The polymerase chain reaction with a single universal primer (UP-PCR) was applied to bacterial strains and mycobacteria isolates alongside conventional methods. A universal protocol of preparation of PCR samples from cultures representing Escherichia coli, Enterobacter aerogenes, Serratia marcescens, Staphylococcus aureus, Streptococcus pyogenes, Klebsiella pneumoniae, Mycobacterium tuberculosis, Mycobacterium bovis, and several non-tuberculous mycobacteria was found to be reproducible and efficient with these organisms. The bands of UP-PCR products observed in an agarose gel after electrophoresis were species-specific and provided an efficient means of distinguishing bacterial species. The applicability of this approach to mycobacteria identification was assessed by comparing the DNA bands obtained for different strains. Three reference strains and 22 clinical isolates of M. tuberculosis and M. bovis produced very similar DNA banding patterns. They comprised a triplet of prominent and several minor fragments within the 200-500 base pair (bp) size range and were the easiest to interpret. The DNA profiles of unrelated mycobacteria clearly differed from each other when subjected to electrophoretic analysis and correlated well with results of culture method. The method provides a real promise of its application in clinical studies as a simple assay for distinguishing between tubercle bacilli.
Bahrmand
A R
AR
Mycobacteriology Department, Pasteur Institute of Iran, Tehran, Iran.
Madani
H
H
Bakayev
V V
VV
Babaei
M H
MH
Samar
G
G
Anashchenko
V
V
D001483:Base Sequence; D017931:DNA Primers; D004269:DNA, Bacterial / Q000235:genetics / Q000302:isolation & purification; D004587:Electrophoresis, Agar Gel; D004926:Escherichia coli / Q000235:genetics; D016680:Genome, Bacterial; D006090:Gram-Negative Bacteria / Q000145:classification / Q000235:genetics*; D006094:Gram-Positive Bacteria / Q000145:classification / Q000235:genetics*; D006801:Humans; D007711:Klebsiella pneumoniae / Q000235:genetics; D008969:Molecular Sequence Data; D009161:Mycobacterium / Q000145:classification* / Q000235:genetics* / Q000302:isolation & purification; D009163:Mycobacterium bovis / Q000235:genetics; D009169:Mycobacterium tuberculosis / Q000235:genetics; D016133:Polymerase Chain Reaction / Q000379:methods*; D015203:Reproducibility of Results; D012706:Serratia marcescens / Q000235:genetics; D013211:Staphylococcus aureus / Q000235:genetics; D013297:Streptococcus pyogenes / Q000235:genetics
D017931:DNA Primers; D004269:DNA, Bacterial