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Identification of a Basic Helix-Loop-Helix-Type Transcription Regulator Gene in Aspergillus oryzae by Systematically Deleting Large Chromosomal Segments

Jin, Feng Jie, Takahashi, Tadashi, Machida, Masayuki, Koyama, Yasuji
Applied and environmental microbiology 2009 v.75 no.18 pp. 5943-5951
Aspergillus oryzae, DNA, basic helix-loop-helix transcription factors, binding proteins, chromosomes, engineering, industrial applications, mutants, phenotype, prediction, regulator genes
We previously developed two methods (loop-out and replacement-type recombination) for generating large-scale chromosomal deletions that can be applied to more effective chromosomal engineering in Aspergillus oryzae. In this study, the replacement-type method is used to systematically delete large chromosomal DNA segments to identify essential and nonessential regions in chromosome 7 (2.93 Mb), which is the smallest A. oryzae chromosome and contains a large number of nonsyntenic blocks. We constructed 12 mutants harboring deletions that spanned 16- to 150-kb segments of chromosome 7 and scored phenotypic changes in the resulting mutants. Among the deletion mutants, strains designated Δ5 and Δ7 displayed clear phenotypic changes involving growth and conidiation. In particular, the Δ5 mutant exhibited vigorous growth and conidiation, potentially beneficial characteristics for certain industrial applications. Further deletion analysis allowed identification of the AO090011000215 gene as the gene responsible for the Δ5 mutant phenotype. The AO090011000215 gene was predicted to encode a helix-loop-helix binding protein belonging to the bHLH family of transcription factors. These results illustrate the potential of the approach for identifying novel functional genes.