title: Development of an activation tagging system for the basidiomycetous medicinal fungus Antrodia cinnamomea.
abstract: This study describes the development of an efficient and reliable activation tagging system for the medicinal fungus Antrodia cinnamomea. For successful Agrobacterium tumefaciens-mediated transformation, different parameters were considered. The Agrobacterium concentration of 5 x 10(8) cfu ml(-1), 1mm acetosyringone, 25-d-old mycelia at 0.2 g ml(-1), and co-culture period of 6d were found to be the most optimal conditions for enhancing the transformation efficiency. The mitotic stability of transferred DNA (T-DNA) was demonstrated by growing eight randomly selected putative transformants in malt extract agar medium for five subcultures. Insertion of T-DNA into the genome of transformants was confirmed by PCR and Southern hybridization. Results showed that 88% of the mutants contained a single T-DNA insertion. Two of the mutants were observed with different triterpenoid profiles compared with the untransformed cultures. Our results suggest a new functional genomics approach to tag the triterpenoid biosynthesis genes in A. cinnamomea.
authors.affiliation: Institute of Biochemical Sciences and Technology, Chaoyang University of Technology, 168 Gifeng E Road, Wufeng, Taichung County 41349, Taiwan.
authors.forename: Emily Chin-Fun
authors.initials: EC
authors.lastname: Chen
authors.forename: Yu-Hsiu
authors.initials: YH
authors.lastname: Su
authors.forename: Selvaraju
authors.initials: S
authors.lastname: Kanagarajan
authors.forename: Dinesh Chandra
authors.initials: DC
authors.lastname: Agrawal
authors.forename: Hsin-Sheng
authors.initials: HS
authors.lastname: Tsay
mesh_terms: D016960:Agrobacterium tumefaciens / Q000235:genetics; D055491:Antrodia / Q000235:genetics* / Q000378:metabolism; D002851:Chromatography, High Pressure Liquid; D004271:DNA, Fungal / Q000235:genetics*; D004277:DNA, Single-Stranded / Q000235:genetics*; D016254:Mutagenesis, Insertional / Q000379:methods*; D009154:Mutation; D014170:Transformation, Genetic; D014315:Triterpenes / Q000032:analysis / Q000378:metabolism
chemical_list: D004271:DNA, Fungal; D004277:DNA, Single-Stranded; D014315:Triterpenes