Visual DNA microarrays for simultaneous detection of Ureaplasma urealyticum and Chlamydia trachomatis coupled with multiplex asymmetrical PCR.
Visual DNA microarrays, based on gold label silver stain (GLSS) and coupled with multiplex asymmetrical PCR, were developed for simultaneous, sensitive and specific detection of Ureaplasma urealyticum and Chlamydia trachomatis. 5'-end-amino-modified oligonucleotides, which were immobilized on glass surface, acted as capturing probes that were designed to bind complementary biotinylated targets DNA. The gold-conjugated streptavidins were introduced to the microarray for specific binding to biotin. The black image of microarray spots, resulting from the precipitation of silver onto nanogold particles bound to streptavidins, were used to detect biotinylated targets DNA visually or with a visible light scanner. Multiplex asymmetrical PCR of U. urealyticum, C. trachomatis and Bacillus subtilis (used as positive control) was performed to prepare abundant biotinylated single-stranded targets DNA, which affected detection efficiency and sensitivity of hybridization on microarray. Plenty of clinical samples of U. urealyticum and C. trachomatis from infected patients were tested using home-made DNA microarrays. For its high sensitivity, good specificity, simplicity, cheapness and speed, the present visual gene-detecting technique has potential applications in clinical fields.
Cao
Xuan
X
College of Life Sciences, Wuhan University, Wuhan 430072, China.
Wang
Ye-Fu
YF
Zhang
Chu-Fu
CF
Gao
Wen-Juan
WJ
D015374:Biosensing Techniques / Q000295:instrumentation* / Q000379:methods; D002692:Chlamydia trachomatis / Q000235:genetics* / Q000302:isolation & purification*; D003124:Colorimetry / Q000295:instrumentation / Q000379:methods; D004269:DNA, Bacterial / Q000032:analysis* / Q000235:genetics; D004867:Equipment Design; D019544:Equipment Failure Analysis; D017403:In Situ Hybridization / Q000295:instrumentation / Q000379:methods; D016133:Polymerase Chain Reaction / Q000295:instrumentation* / Q000379:methods; D016990:Ureaplasma urealyticum / Q000235:genetics* / Q000302:isolation & purification*
D004269:DNA, Bacterial