Improvement of kojic acid production in Aspergillus oryzae AR-47 mutant strain by combined mutagenesis.

Kojic acid is a kind of secondary metabolites, whose biosynthesis pathway remains unclear to date. It is produced industrially by microbial fermentation, and thus, mutagenesis breeding still plays a vital role for obtaining strains with high kojic acid production. The starting strain KA-11 isolated from mildewed fruits was identified as Aspergillus oryzae and then subjected to a combined mutagenesis program including microwave mutagenesis, UV irradiation, heat-LiCl, atmospheric, and room temperature plasma (ARTP). The kojic acid production was increased by 47.0%, 87.1%, 126.2%, and 292.3% compared with the starting strain KA-11 after each mutagenesis stage. A mutant strain AR-47 with kojic acid production of 96.5 g/L in flask-shake fermentation was finally obtained. The fermentation time also decreased from 7 to 5 days. Real-time quantitative PCR was used to determine the transcriptional expression levels of genes that may be relevant to kojic acid biosynthesis, including kojA, kojR, kojT, AO090113000141, AO090113000143, AO090113000145, nrtA, and laeA. The results showed that the transcriptional expression levels of all these genes in high yield strain AR-47 had increased compared with the starting strain KA-11.