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A screening method for the detection of the 35S promoter and the nopaline synthase terminator in genetically modified organisms in a real-time multiplex polymerase chain reaction using high-resolution melting-curve analysis.
Akiyama, Hiroshi; Nakamura, Fumi; Yamada, Chihiro; Nakamura, Kosuke; Nakajima, Osamu; Kawakami, Hiroshi; Harikai, Naoki; Furui, Satoshi; Kitta, Kazumi; Teshima, Reiko.
Afiliação
  • Akiyama H; National Institute of Health Sciences, 1-18-1 Kamiyoga, Setagaya-ku, Tokyo 158-8501, Japan. akiyama@nihs.go.jp
Biol Pharm Bull ; 32(11): 1824-9, 2009 Nov.
Article em En | MEDLINE | ID: mdl-19881291
To screen for unauthorized genetically modified organisms (GMO) in the various crops, we developed a multiplex real-time polymerase chain reaction high-resolution melting-curve analysis method for the simultaneous qualitative detection of 35S promoter sequence of cauliflower mosaic virus (35SP) and the nopaline synthase terminator (NOST) in several crops. We selected suitable primer sets for the simultaneous detection of 35SP and NOST and designed the primer set for the detection of spiked ColE1 plasmid to evaluate the validity of the polymerase chain reaction (PCR) analyses. In addition, we optimized the multiplex PCR conditions using the designed primer sets and EvaGreen as an intercalating dye. The contamination of unauthorized GMO with single copy similar to NK603 maize can be detected as low as 0.1% in a maize sample. Furthermore, we showed that the present method would be applicable in identifying GMO in various crops and foods like authorized GM soybean, authorized GM potato, the biscuit which is contaminated with GM soybeans and the rice which is contaminated with unauthorized GM rice. We consider this method to be a simple and reliable assay for screening for unauthorized GMO in crops and the processing food products.
Assuntos
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Coleções: 01-internacional Base de dados: MEDLINE Assunto principal: Reação em Cadeia da Polimerase / Regiões Promotoras Genéticas / Regiões Terminadoras Genéticas / Plantas Geneticamente Modificadas / Aminoácido Oxirredutases Tipo de estudo: Diagnostic_studies / Qualitative_research / Screening_studies Idioma: En Revista: Biol Pharm Bull Assunto da revista: BIOQUIMICA / FARMACOLOGIA Ano de publicação: 2009 Tipo de documento: Article País de afiliação: Japão
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Coleções: 01-internacional Base de dados: MEDLINE Assunto principal: Reação em Cadeia da Polimerase / Regiões Promotoras Genéticas / Regiões Terminadoras Genéticas / Plantas Geneticamente Modificadas / Aminoácido Oxirredutases Tipo de estudo: Diagnostic_studies / Qualitative_research / Screening_studies Idioma: En Revista: Biol Pharm Bull Assunto da revista: BIOQUIMICA / FARMACOLOGIA Ano de publicação: 2009 Tipo de documento: Article País de afiliação: Japão