Construction and characterization of a DNA probe for distinguishing strains of Aspergillus flavus.

Abstract

Repetitive DNA sequences have proven useful and reliable characters in evaluating genetic relatedness of strains at different levels of taxonomic classification. A DNA probe was constructed to distinguish among strains of Aspergillus flavus by DNA fingerprinting techniques. Chromosomal DNA of A. flavus var. flavus NRRL 6541 was partially digested with EcoRI and ligated to a Lambda Dash bacteriophage vector. Four lambda clones were identified which displayed multiple and distinct bands when hybridized with chromosomal DNA from seven strains of A. flavus var. flavus digested with either EcoRI or PstI. One of these clones was chosen for further analysis and was subcloned into pUC19. The subclone, pAF28, contained a 6.2-kb chromosomal DNA insert and was able to distinguish among strains characterized by K. E. Papa (Mycologia 78:98-101, 1986) as belonging to 22 different vegetative compatibility groups. The subclone identified unique banding patterns when hybridized to genomic DNA digested with PstI. The cloned probe may be species specific as it hybridized with the DNA of all isolates of A. flavus tested in addition to strains recognized as varieties of A. flavus (e.g., A. flavus var. oryzae, A. flavus var. parasiticus, and A. flavus var. sojae). pAF28 hybridized to a single band on a Southern blot with Aspergillus nomius DNA but did not hybridize with the DNA of other fungal species tested including Aspergillus ochraceus, Aspergillus auricomus, Aspergillus alliaceus, Fusarium moniliforme, and Penicillium thomii.