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Microfluidic device for integrated restriction digestion reaction and resulting DNA fragment analysis
Author(s) -
Xie Hua,
Li Bowei,
Zhong Runtao,
Qin Jianhua,
Zhu Yisheng,
Lin Bingcheng
Publication year - 2008
Publication title -
electrophoresis
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.666
H-Index - 158
eISSN - 1522-2683
pISSN - 0173-0835
DOI - 10.1002/elps.200800490
Subject(s) - microfluidics , electrophoresis , chromatography , analyte , materials science , restriction digest , restriction enzyme , capillary electrophoresis , bamhi , chemistry , analytical chemistry (journal) , nanotechnology , dna , biochemistry
A microfluidic system combining temperature‐controlled reactor, analyte delivery, chip electrophoresis (CE) separation, and fluorescence detection was developed, in which the heaters, resistance temperature detectors, enzymatic reactors, CE channels, and pneumatic valves/pumps were integrated onto a single glass–PDMS chip. The microdevice was used to perform the digestion reaction, followed by on‐line electrophoresis separation and detection of the resulting fragments with endonuclease Bam HI and Fok I as models. Pneumatic valves/pumps served not only for isolating the reaction region from the separation medium to prevent contamination, but also for delivering and quantitatively diluting the fluid from the reaction chamber to the CE section. Thus enzymatic reaction and electrophoresis separation could be insulated and connected as needed. A dynamic coating procedure with the use of PVP and mannitol was firstly adopted for glass–PDMS hybrid chip‐based DNA separations, leading to an improved separation efficiency with reproducible migration time and theoretical plates. The expected 263‐ and 287‐bp digestion products of Bam HI and Fok I were definitely verified by the size‐based electrophoretic separation and detection. The whole integrated reaction‐CE system can be manipulated in a simple manner with good reproducibility, which is expected to be applied in other on‐line analysis of various biochemical reactions.