Transposon mutagenesis using a resident DNA transposon Crawler in Aspergillus oryzae

Hironobu Ogasawara 1, Tsutomu Satoh 2, Hiroshi Konno 2, Yoji Hata 3, Saori Takahashi1, and Katsuya Gomi 4

Author address: 

Akita Res. Inst. Food and Brewing, Akita, Japan. 2 Akita Konno Co. Ltd., Akita, Japan, 3Research Institute, Gekkeikan Sake Co. Ltd., Kyoto, Japan. 4 Graduate School of Agricultural Science, Tohoku University, Sendai, Japan. E-mail:[email protected]

Abstract: 

An active DNA transposon Crawler isolated from the genome of industrially important fungus Aspergillus oryzae transposes under extreme stress conditions. The DNA sequencing surveys revealed that the Crawler element is widely distributed among A. oryzae and A. sojae strains, which are commonly used in Japanese traditional fermentation manufacturing. In the present study, we analyzed the relationship between various stress stimuli and inhibition of cryptic splicing of the Crawler mRNA by qRT-PCR to enhance the frequency of Crawler-mediated mutagenesis in an A. oryzae industrial strain, AOK139. Under the optimized stress conditions, in which conidiospores were treated in 20mM CuSO 4 or 52 °C for 6hr, various phenotypic mutants different from the parent strain were isolated. Those exhibited white color in conidiospores, less number of spores formed, shortened aerial hyphae, thin colony mat and so on. DNA sequencing analyses of a white conidia mutant revealed that Crawler was newly inserted within a coding region of wA gene encoding polyketide synthetase, which resulted in wA deficiency. The insertion occurred also at a TA site with duplication according to the manner of Crawler transposition. These results suggested that transposon mutagenesis using active Crawler is potentially valuable to improve characteristics of A. oryzae industrial strains.
2010

abstract No: 

28

Full conference title: 

7th International Aspergillus Meeting
    • Asperfest 7 (2010)