DNA hybridization detection in a microfluidic channel using two fluorescently labelled nucleic acid probes.
A conceptually new technique for fast DNA detection has been developed. Here, we report a fast and sensitive online fluorescence resonance energy transfer (FRET) detection technique for label-free target DNA. This method is based on changes in the FRET signal resulting from the sequence-specific hybridization between two fluorescently labelled nucleic acid probes and target DNA in a PDMS microfluidic channel. Confocal laser-induced microscopy has been used for the detection of fluorescence signal changes. In the present study, DNA hybridizations could be detected without PCR amplification because the sensitivity of confocal laser-induced fluorescence detection is very high. Two probe DNA oligomers (5'-CTGAT TAGAG AGAGAA-TAMRA-3' and 5'-TET-ATGTC TGAGC TGCAGG-3') and target DNA (3'-GACTA ATCTC TCTCT TACAG GCACT ACAGA CTCGA CGTCC-5') were introduced into the channel by a microsyringe pump, and they were efficiently mixed by passing through the alligator teeth-shaped PDMS microfluidic channel. Here, the nucleic acid probes were terminally labelled with the fluorescent dyes, tetrafluororescein (TET) and tetramethyl-6-carboxyrhodamine (TAMRA), respectively. According to our confocal fluorescence measurements, the limit of detection of the target DNA is estimated to be 1.0 x 10(-6) to 1.0 x 10(-7)M. Our result demonstrates that this analytical technique is a promising diagnostic tool that can be applied to the real-time analysis of DNA targets in the solution phase.
Chen
Lingxin
L
Department of Applied Chemistry, Hanyang University, Sa-1-dong 127, Ansan, Kyunggi-do 426-791, Republic of Korea.
Lee
Sangyeop
S
Lee
Moonkwon
M
Lim
Chaesung
C
Choo
Jaebum
J
Park
Joong Yull
JY
Lee
Sanghoon
S
Joo
Sang-Woo
SW
Lee
Kyeong-Hee
KH
Choi
Young-Wook
YW
D004247:DNA / Q000032:analysis* / Q000235:genetics*; D015342:DNA Probes / Q000737:chemistry* / Q000235:genetics*; D004867:Equipment Design; D019544:Equipment Failure Analysis; D017404:In Situ Hybridization, Fluorescence / Q000295:instrumentation* / Q000379:methods; D046210:Microfluidic Analytical Techniques / Q000295:instrumentation*; D013194:Staining and Labeling / Q000379:methods
D015342:DNA Probes; D004247:DNA