Περίληψη:
A microfabricated, inexpensive, reusable glass capillary electrophoresis
chip and a laser-induced fluorescence system were developed in-house for
the rapid DNA-based analysis of genetically modified organisms (GMOs).
The 35S promoter sequence of cauliflower mosaic virus and the terminator
of the nopaline synthase (NOS) gene from Agrobacterium tumefaciens were
both detected since they are present in most genetically modified
organisms. The detection of genetically modified soybean in the presence
of unaltered soybean was chosen as a model. Lectin, a plant-specific
gene, was also detected for confirmation of the integrity of extracted
DNA. The chip was composed of two glass plates, each 25 x 76 mm,
thermally bonded together to form a closed structure. Photomasks with
cross-topology were prepared rapidly by using polymeric material instead
of chrome plates. The widths of the injection and separation channels
were 30 and 70 pm, respectively, the effective separation length 4.5 cm.
The glass slide was etched to a depth of 30 mum for both the injection
and separation channel. The cost of the chip was less than 1 $ and
required 2 days for photomask preparation and microfabrication. The
separation and detection of polymerase chain reaction-amplified NOS,
35S, and lectin sequences (180, 195, and 181 bp, respectively) was
completed in less than 60 s. As low as 0.1% GMO content was detectable
by the proposed system after 35 and 40 amplification cycles for 35S and
NOS, respectively, using 25 ng of extracted DNA as starting material.
This corresponds to only 20 genome copies of genetically modified
soybean.
Συγγραφείς:
Obeid, PJ
Christopoulos, TK
Ioannou, PC
