title: Comparison and evaluation of two diagnostic methods for detection of npt II and GUS genes in Nicotiana tabacum.
abstract: To diminish the time required for some diagnostic assays including polymerase chain reaction (PCR), loop-mediated isothermal amplification (LAMP) and also a visual detection protocol on the basis of npt II and GUS genes in transgenic tobacco plants were used. Agrobacterium tumefaciens-mediated transformation of Nicotiana tabacum leaf discs was performed with plant transformation vector of pBI 121. From kanamycin-resistant plants selected by their antibiotic resistance, four plants were selected for DNA isolation. Presence of the transgene was confirmed in the transformants by PCR and LAMP. In this regard, all LAMP and PCR primers were designed on the basis of the gene sequences of npt II and GUS. The LAMP assay was applied for direct detection of gene marker from plant samples without DNA extraction steps (direct LAMP assay). Also, a novel colorimetric LAMP assay for rapid and easy detection of npt II and GUS genes was developed here, its potential compared with PCR assay. The LAMP method, on the whole, had the following advantages over the PCR method: easy detection, high sensitivity, high efficiency, simple manipulation, safety, low cost, and user friendly.
authors.affiliation: Young Researchers and Elite Club, North Tehran Branch, Islamic Azad University, Tehran, Iran, mohammadaminalmasi136301@gmail.com.
authors.forename: Mohammad Amin
authors.initials: MA
authors.lastname: Almasi
authors.forename: Mehdi
authors.initials: M
authors.lastname: Aghapour-Ojaghkandi
authors.forename: Khadijeh
authors.initials: K
authors.lastname: Bagheri
authors.forename: Mohammadreza
authors.initials: M
authors.lastname: Ghazvini
authors.forename: Seyed Mohammad
authors.initials: SM
authors.lastname: Hosseyni-Dehabadi
mesh_terms: D016960:Agrobacterium tumefaciens / Q000235:genetics*; D005822:Genetic Vectors; D005966:Glucuronidase / Q000235:genetics / Q000302:isolation & purification*; D019868:Kanamycin Kinase / Q000235:genetics / Q000302:isolation & purification*; D018515:Plant Leaves / Q000235:genetics; D030821:Plants, Genetically Modified; D016133:Polymerase Chain Reaction; D014026:Nicotiana / Q000201:enzymology / Q000235:genetics*
chemical_list: D019868:Kanamycin Kinase; D005966:Glucuronidase