Genetic Approach to Target Putative Pks Genes in <i>Aspergillus</i> Section Nigri Species Producing Ochratoxin A

Abstract

Using degenerated primers (LC1–LC2c) and two novel primer pairs, namely (KSLB–LC6) for Aspergillus niger and (AFl1F–LC2) for Aspergillus tubingensis, created for the acyl transferase (AT) and the KS sequences of fungal PKSs genes, a 700 pb PCR-derived DNA fragment was isolated from Aspergillus carbonarius, Aspergillus niger, and Aspergillus tubingensis. Testing was performed on DNA from most of the black Aspergillus species currently known to exist. This article describes the identification and characterisation of a portion of a novel putative OTA-polyketide synthase gene in A. tubingensis “AT Pks,” A. niger “AN Pks,” and A. carbonarius “AC Pks”. Phylogenetic methods were used to align and evaluate the sequences. The study’s primers demonstrated broad application, and several Aspergillus species from the section Nigri, particularly A. niger and A. tubingensis, were amplified satisfactorily. Predicted amino acid sequences known as “AC Pks” showed 66–81% similarity to several polyketide synthase genes, whereas “AN Pks” and “AT Pks” showed 68–71% and 81–97% similarity, respectively. The AT and KS sequences were linked to PKSs engaged in various mycotoxin production routes, including ochratoxin A, and they seemed to be specific for a specific kind of fungal PKSs. The sequences that have been reported in this paper are particularly useful in finding new fungal PKS gene clusters.

Keywords

• PKS genes
• OTA biosynthesis
• <i>Aspergillus</i> Nigri
• sequence analysis