Kojic acid biosynthesis in Aspergillus oryzae is regulated by a Zn(II)(2)Cys(6) transcriptional activator and induced by kojic acid at the transcriptional level.

A gene encoding the Zn(II)(2)Cys(6) transcriptional factor is clustered with two genes involved in biosynthesis of a secondary metabolite, kojic acid (KA), in Aspergillus oryzae. We determined that the gene was essential for KA production and the transcriptional activation of KA biosynthetic genes, which were triggered by the addition of KA.

Penicillin biosynthesis in Aspergillus oryzae and its overproduction by genetic engineering.

Aspergillus oryzae penicillin biosynthetic genes were clustered. The penicillin production was positively regulated by VeA, a global gene regulator required for transcriptional expression of the penicillin biosynthetic genes. Overexpression of the biosynthetic genes by a strong promoter yielded a greater than 100-fold increase in penicillin production.

A novel vector for positive selection of inserts harboring an open reading frame by translational coupling.

We have developed a novel vector pTCS, as a tool for efficient selection of open reading frame (ORF)-containing inserts. In pTCS clones containing an insert with an ORF a downstream marker gene (immE3, conferring resistance to colicin) is activated via translational coupling with the insert, and transformed cells can then be selected by exposure to colicin E3. Our method differs from previous methods in that the marker gene is activated without protein fusion, and that selection occurs irrespective of the reading frame of the insert.

Analysis of expressed sequence tags from the fungus Aspergillus oryzae cultured under different conditions.

We performed random sequencing of cDNAs from nine biologically or industrially important cultures of the industrially valuable fungus Aspergillus oryzae to obtain expressed sequence tags (ESTs). Consequently, 21 446 raw ESTs were accumulated and subsequently assembled to 7589 non-redundant consensus sequences (contigs). Among all contigs, 5491 (72.

Construction of a positive selection marker by a lethal gene with the amber stop codon(s) regulator.

A novel positive selection marker for Escherichia coli transformation was developed. The marker consisted of a DNA fragment encoding the C-terminal ribonuclease domain (CRD) of colicin E3 (colE3) and one or more amber stop codons between the initiation codon and the E3-CRD coding sequence. The toxicity of the marker was controlled by the suppressor activity the host cells possessed.