Gene silencing by RNA interference in the koji mold Aspergillus oryzae.
We found the orthologous genes required for RNA interference (RNAi) in the Aspergillus oryzae genome database, and constructed a set of tools for gene silencing using RNAi in A. oryzae. This system utilizes compatible restriction enzyme sites so that only a single target gene fragment is required to create the hairpin RNA cassette. For ease of handling, we also separated the construction of the hairpin RNA cassette for the target gene from its subsequent introduction into the expression vector. Using the brlA gene as a target for RNAi, we detected decreased mRNA levels and a delayed conidiation phenotype in the transformants. Furthermore, even though A. oryzae possesses three copies of the alpha-amylase gene, a single copy of an alpha-amylase RNAi construct was sufficient to downregulate the mRNA levels and decrease the enzymatic activity to 10% of control levels. Gene silencing by RNAi should provide a powerful genetic tool for post-genomic studies of the industrially important fungus A. oryzae.
Yamada
Osamu
O
National Research Institute of Brewing, Japan. yamada@nrib.go.jp
Ikeda
Ryoko
R
Ohkita
Yuka
Y
Hayashi
Risa
R
Sakamoto
Kazutoshi
K
Akita
Osamu
O
D001236:Aspergillus oryzae / Q000235:genetics* / Q000378:metabolism; D005822:Genetic Vectors / Q000235:genetics; D016681:Genome, Fungal / Q000235:genetics; D034622:RNA Interference; D014157:Transcription Factors / Q000235:genetics / Q000378:metabolism; D000516:alpha-Amylases / Q000235:genetics / Q000378:metabolism
D014157:Transcription Factors; D000516:alpha-Amylases