Isolation of industrial strains of Aspergillus oryzae lacking ferrichrysin by disruption of the dffA gene.
Based on studies using laboratory strains, the efficiency of gene disruption in Aspergillus oryzae, commonly known as koji mold, is low; thus, gene disruption has rarely been applied to the breeding of koji mold. To evaluate the efficiency of gene disruption in industrial strains of A. oryzae, we produced ferrichrysin biosynthesis gene (dffA) disruptants using three different industrial strains as hosts. PCR analysis of 438 pyrithiamine-resistant transformants showed dffA gene disruption efficiency of 42.9%-64.1%, which is much higher than previously reported. Analysis of the physiological characteristics of the disruptants indicated that dffA gene disruption results in hypersensitivity to hydrogen peroxide. To investigate the industrial characteristics of dffA gene disruptants, two strains were used to make rice koji and their properties were compared to those of the host strains. No differences were found between the dffA gene disruptants and the host strains, except that the disruptants did not produce ferrichrysin. Thus, this gene disruption technique is much more effective than conventional mutagenesis for A. oryzae breeding.
Watanabe
Hisayuki
H
Iwate Biotechnology Research Center, 22-174-4 Narita, Kitakami, Iwate, Japan.
Hatakeyama
Makoto
M
Sakurai
Hiroshi
H
Uchimiya
Hirofumi
H
Sato
Toshitsugu
T
D001236:Aspergillus oryzae / Q000235:genetics*; D001709:Biotechnology / Q000379:methods*; D015139:Blotting, Southern; D017931:DNA Primers / Q000737:chemistry; D005291:Ferrichrome / Q000031:analogs & derivatives* / Q000737:chemistry / Q000378:metabolism; D005822:Genetic Vectors; D006861:Hydrogen Peroxide / Q000737:chemistry; D007218:Industrial Microbiology / Q000379:methods*; D008957:Models, Genetic; D016296:Mutagenesis; D012275:Oryza; D010957:Plasmids / Q000378:metabolism; D016133:Polymerase Chain Reaction; D011745:Pyrithiamine / Q000737:chemistry
D017931:DNA Primers; C513415:ferrichrysin; D005291:Ferrichrome; D011745:Pyrithiamine; D006861:Hydrogen Peroxide