title: Im"plant"ing of Mammalian Glycosyltransferase Gene into Plant Suspension-Cultured Cells Using Agrobacterium-Mediated Transformation.
abstract: Enzymatic activity assay of exogenous glycosyltransferase (GT) and glycosylhydrolase (GH) expressed in plants is an important analysis for determination of the expression of the gene of interest. However, generations and establishment of in planta transgenic lines are time-consuming. Furthermore, the expression levels and the activities of the exogenous GTs and GHs are quite low and weak, the radiolabeled donor substrate had to be used to analyze the enzymatic activity. Here, we describe a protocol for the generation of transgenic plants using suspension-cultured cells and a high sensitive assay for GT, especially β1,4-galactosyltransferase, using microsomal fraction from plant cells and fluorescent-labeled sugar chains as an acceptor substrate. This method enables less-time-consuming preparation of stable transgenic plants, non-radiolabeled, high-throughput detail analysis which includes mass spectrometric analysis and exo-glycosidase digestions.
authors.affiliation: The International Center for Biotechnology, Osaka University, Osaka, Japan.
authors.forename: Hiroyuki
authors.initials: H
authors.lastname: Kajiura
authors.forename: Kazuhito
authors.initials: K
authors.lastname: Fujiyama
mesh_terms: D060054:Agrobacterium / Q000235:genetics*; D000818:Animals; D018929:Cell Culture Techniques; D002460:Cell Line; D016695:Glycosyltransferases / Q000235:genetics*; D008322:Mammals / Q000235:genetics*; D059828:Plant Cells / Q000378:metabolism*; D010940:Plant Proteins / Q000235:genetics*; D030821:Plants, Genetically Modified / Q000235:genetics*; D014170:Transformation, Genetic / Q000235:genetics*
chemical_list: D010940:Plant Proteins; D016695:Glycosyltransferases