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Title

Simultaneous detection of genetically modified organisms by multiplex ligation-dependent genome amplification and capillary gel electrophoresis with laser-induced fluorescence

AuthorsGarcía-Cañas, Virginia ; Mondello, Monica; Cifuentes, Alejandro
KeywordsCGE-LIF
DNA amplification
Food analysis
Genetically modified organisms
Transgenic maize
Issue DateJul-2010
PublisherWiley-VCH
CitationElectrophoresis 31(13): 2249-2259 (2010)
AbstractIn this work, an innovative method useful to simultaneously analyze multiple genetically modified organisms is described. The developed method consists in the combination of multiplex ligation-dependent genome dependent amplification (MLGA) with CGE and LIF detection using bare-fused silica capillaries. The MLGA process is based on oligonucleotide constructs, formed by a universal sequence (vector) and long specific oligonucleotides (selectors) that facilitate the circularization of specific DNA target regions. Subsequently, the circularized target sequences are simultaneously amplified with the same couple of primers and analyzed by CGE-LIF using a bare-fused silica capillary and a run electrolyte containing 2-hydroxyethyl cellulose acting as both sieving matrix and dynamic capillary coating. CGE-LIF is shown to be very useful and informative for optimizing MLGA parameters such as annealing temperature, number of ligation cycles, and selector probes concentration. We demonstrate the specificity of the method in detecting the presence of transgenic DNA in certified reference and raw commercial samples. The method developed is sensitive and allows the simultaneous detection in a single run of percentages of transgenic maize as low as 1% of GA21, 1% of MON863, and 1% of MON810 in maize samples with signal-to-noise ratios for the corresponding DNA peaks of 15, 12, and 26, respectively. These results demonstrate, to our knowledge for the first time, the great possibilities of MLGA techniques for genetically modified organisms analysis.
Description11 páginas, 5 figuras, 2 tablas.-- El pdf del artículo es la versión revisada de autor.
Publisher version (URL)http://dx.doi.org/10.1002/elps.201000078
URIhttp://hdl.handle.net/10261/50320
DOI10.1002/elps.201000078
ISSN0173-0835
E-ISSN1522-2683
Appears in Collections:(IFI) Artículos
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