Simultaneous strand displacement amplification and fluorescence polarization detection of Chlamydia trachomatis DNA.
Strand displacement amplification (SDA) is an isothermal DNA amplification technology that uses a restriction enzyme and polymerase. We have developed a target-specific method which allows simultaneous SDA and detection in a homogeneous format. This is accomplished by including a detector oligodeoxynucleotide labeled with 5-(4,6-dichlorotriazin-2-yl)amino fluorescein in the SDA reaction. Fluorescence polarization is used to monitor hybridization of the detector probe to the amplification product as it rises in concentration during SDA. We have demonstrated real-time SDA detection for the cryptic plasmid of Chlamydia trachomatis with high sensitivity in only 30 min.
Spears
P A
PA
Becton Dickinson Research Center, Research Triangle Park, North Carolina 27709, USA.
Linn
C P
CP
Woodard
D L
DL
Walker
G T
GT
D001483:Base Sequence; D002690:Chlamydia Infections / Q000175:diagnosis; D002692:Chlamydia trachomatis / Q000235:genetics* / Q000302:isolation & purification*; D004269:DNA, Bacterial / Q000032:analysis* / Q000235:genetics*; D005069:Evaluation Studies as Topic; D005260:Female; D019793:Fluorescein; D005452:Fluoresceins; D005454:Fluorescence Polarization / Q000379:methods* / Q000706:statistics & numerical data; D005456:Fluorescent Dyes; D006801:Humans; D008297:Male; D021141:Nucleic Acid Amplification Techniques; D015345:Oligonucleotide Probes / Q000235:genetics; D012680:Sensitivity and Specificity; D013997:Time Factors
D004269:DNA, Bacterial; D005452:Fluoresceins; D005456:Fluorescent Dyes; D015345:Oligonucleotide Probes; C039083:5-((4,6-dichloro-1,3,5-triazin-2-yl)amino)fluorescein; D019793:Fluorescein