Microbead-based rolling circle amplification in a microchip for sensitive DNA detection.
The sensitive detection and quantification of DNA targets in the food industry and in environmental and clinical settings are issues of utmost importance in ensuring contamination-free food, monitoring the environment, and battling disease. Selective probes coupled with powerful amplification techniques are therefore of major interest. In this study, we set out to create an integrated microchemical chip that benefits from microfluidic chip technology in terms of sensitivity and a strong detection methodology provided jointly by padlock probes and rolling circle amplification (RCA). Here, we have integrated padlock probes and RCA into a microchip. The chip uses solid phase capture in a microchannel to enable washing cycles and decrease analytical area, and employs on-bead RCA for single-molecule amplification and detection. We investigated the effects of reagent concentration and amount of padlock probes, and demonstrated the feasibility of detecting Salmonella.
Sato
Kae
K
Center for Nanobio Integration, The University of Tokyo, 7-3-1 Hongo, Bunkyo, Tokyo 113-8656, Japan.
Tachihara
Atsuki
A
Renberg
Björn
B
Mawatari
Kazuma
K
Sato
Kiichi
K
Tanaka
Yuki
Y
Jarvius
Jonas
J
Nilsson
Mats
M
Kitamori
Takehiko
T
D001483:Base Sequence; D004247:DNA / Q000032:analysis* / Q000235:genetics*; D004269:DNA, Bacterial / Q000032:analysis / Q000235:genetics; D046208:Microchip Analytical Procedures / Q000379:methods*; D008863:Microspheres; D008904:Miniaturization; D021141:Nucleic Acid Amplification Techniques / Q000295:instrumentation / Q000379:methods*; D012475:Salmonella / Q000235:genetics / Q000302:isolation & purification; D013997:Time Factors
D004269:DNA, Bacterial; D004247:DNA