Bt11 event detection by real-time PCR: single-laboratory validation, comparison of DNA extraction and quantification techniques and application

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dc.creatorCarolina Sheng Whei Miaw
dc.creatorEmanuel Novaes Vasconcelos
dc.creatorNilson César Castanheira Guimarães
dc.creatorScheilla Vitorino Carvalho de Souza
dc.date.accessioned2022-05-06T22:08:07Z
dc.date.accessioned2025-09-09T00:30:10Z
dc.date.available2022-05-06T22:08:07Z
dc.date.issued2017
dc.description.sponsorshipCNPq - Conselho Nacional de Desenvolvimento Científico e Tecnológico
dc.format.mimetypepdf
dc.identifier.doi10.3920/qas2016.0974
dc.identifier.issn1757-8361
dc.identifier.urihttps://hdl.handle.net/1843/41441
dc.languageeng
dc.publisherUniversidade Federal de Minas Gerais
dc.relation.ispartofQuality Assurance and Safety of Crops & Foods
dc.rightsAcesso Aberto
dc.subjectTecnologia de alimentos
dc.subjectMilho
dc.subjectDNA
dc.subjectOrganismos geneticamente modificados
dc.subject.otherqualitative validation
dc.subject.othergenetically modified organisms
dc.subject.otherBt11 maize
dc.subject.otherreal time PCR
dc.subject.otherDNA extraction
dc.subject.otherDNA quantification
dc.titleBt11 event detection by real-time PCR: single-laboratory validation, comparison of DNA extraction and quantification techniques and application
dc.typeArtigo de periódico
local.citation.epage412
local.citation.issue4
local.citation.spage401
local.citation.volume9
local.description.resumoThe detection of genetically modified organisms (GMO) by real-time polymerase chain reaction (PCR) is recommended due to its effectiveness in GMO analysis. A complete in-house validation method was applied to the detection of Bt11 events by real-time PCR. A full factorial design was used to compare DNA extraction (cetyltrimethyl ammonium bromide; CTAB, and NucleoSpin® Plant II Kit) and DNA quantification techniques (conventional GENESYS™ 10S UV-Vis spectrophotometer and confined drop-based NANOVUE™ Plus spectrophotometer). In the validation, various levels (0.0007 to 0.0315%) of Bt11 maize were formulated with blank maize and certified Bt11 reference material. A false-positive rate of 0% was obtained for blank samples, which corresponded to selectivity and reliability rates of 100%. The false-negative rate varied from 0 to 83.3%, consistent with sensitivity and reliability rates ranging from 16.7 to 100%. The Bt11 level that presented 100% positive results was 0.0315%, which indicated the sensitivity of the method. Non-linear models were used to estimate the region of unreliability and to calculate the detection limit of 0.014%. Accordance and concordance values of 1.0 were obtained for the 0.0315% level, which indicated method standardisation. Selectivity in the presence of interference was confirmed by the detection of Bt11 maize in the presence of other events. The method was considered robust for different DNA extraction and DNA quantification techniques. Higher DNA concentration values were obtained using CTAB. The absorbance ratio of A260/A230 was negatively influenced by quantification using a conventional spectrophotometer. Both DNA extraction techniques gave values of A260/A280 higher than 1.7, which indicated DNA of great purity. This validated method was applied to routine samples.
local.publisher.countryBrasil
local.publisher.departmentFAR - DEPARTAMENTO DE ALIMENTOS
local.publisher.initialsUFMG
local.url.externahttps://qascf.com/index.php/qas/article/view/115

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