title: Stable transformants of the azaphilone pigment-producing Monascus purpureus obtained by protoplast transformation and Agrobacterium-mediated DNA transfer.
abstract: The high-level pigment-producing Monascus strain IBCC1 was characterized by random amplification of polymorphic DNA as M. purpureus. This technique allowed us to distinguish between M. purpureus and M. ruber strains. Transformation of Monascus species has not been previously reported. Protoplast formation and regeneration from M. purpureus IBCC1 was optimized by modification of growth media, lytic enzyme mixture, osmotic stabilizer and regeneration media. Of the Monascus transformants, 60% were found to be mitotically stable and retained the plasmid inserted in the chromosome after repeated sporulation cycles. Additionally, an Agrobacterium-mediated DNA transfer system was developed. The transformants obtained by Agrobacterium-mediated DNA transfer remained fully stable (98%) after four sporulation rounds and showed bands of hybridization corresponding to integration of the plasmid in different sites of the genome. The green fluorescent protein marker was well expressed in the M. purpureus transformants. The development of transformation systems is a basic tool for advanced genetic manipulation of the natural pigment producers, M. purpureus and M. ruber.
authors.affiliation: Instituto de Biotecnología de León, Parque Científico de León, Avenida del Real 1, 24006, León, Spain.
authors.forename: Sonia
authors.initials: S
authors.lastname: Campoy
authors.forename: Flor
authors.initials: F
authors.lastname: Pérez
authors.forename: Juan F
authors.initials: JF
authors.lastname: Martín
authors.forename: Santiago
authors.initials: S
authors.lastname: Gutiérrez
authors.forename: Paloma
authors.initials: P
authors.lastname: Liras
mesh_terms: D004269:DNA, Bacterial / Q000235:genetics; D018014:Gene Transfer Techniques; D005822:Genetic Vectors / Q000235:genetics; D049452:Green Fluorescent Proteins; D008164:Luminescent Proteins / Q000378:metabolism; D032741:Monascus / Q000235:genetics* / Q000302:isolation & purification / Q000378:metabolism; D010860:Pigments, Biological / Q000378:metabolism; D010957:Plasmids / Q000235:genetics; D011523:Protoplasts / Q000737:chemistry; D012231:Rhizobium / Q000235:genetics*; D014170:Transformation, Genetic
chemical_list: D004269:DNA, Bacterial; D008164:Luminescent Proteins; D010860:Pigments, Biological; D049452:Green Fluorescent Proteins