A beta-glucuronidase reporter gene construct for monitoring aflatoxin biosynthesis in Aspergillus flavus

Aflatoxins are toxic and carcinogenic secondary metabolites produced by the fungi Aspergillus flavus and A. parasiticus. Current research is directed at the elimination of these compounds in important food sources. The objective of this research was to develop a method to study the induction and reg...

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Bibliographic Details
Published in:Applied and Environmental Microbiology 1995-07, Vol.61 (7), p.2482-2486
Main Authors: Flaherty, J.E. (North Carolina State University, Raleigh, NC.), Weaver, M.A, Payne, G.A, Woloshuk, C.P
Format: Article
Language:English
Subjects:
ADN RECOMBINADO
ADN RECOMBINE
AFLATOXINAS
AFLATOXINE
Aflatoxins - biosynthesis
AMANDE DE LA GRAINE
ASPERGILLUS FLAVUS
Aspergillus flavus - genetics
Aspergillus flavus - metabolism
Aspergillus parasiticus
Bacteria
Bacteriology
Biochemistry
Biological and medical sciences
BIOSINTESIS
BIOSYNTHESE
EXPRESION GENICA
EXPRESSION DES GENES
Fundamental and applied biological sciences. Psychology
GENE
GENES
Genes, Reporter
GERMEN
GLICOSIDASAS
Glucuronidase - genetics
GLYCOSIDASE
Microbiology
Mycology
Pathogenicity, host-agent relations, miscellaneous strains, epidemiology
TRANSFORMACION GENETICA
TRANSFORMATION GENETIQUE
ZEA MAYS
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Summary:Aflatoxins are toxic and carcinogenic secondary metabolites produced by the fungi Aspergillus flavus and A. parasiticus. Current research is directed at the elimination of these compounds in important food sources. The objective of this research was to develop a method to study the induction and regulation of aflatoxin biosynthesis by examining the expression of one aflatoxin pathway gene, ver1. The promoter region of ver1 was fused to the beta-glucuronidase (GUS) gene (uidA) from Escherichia coli to form the reporter construct, GAP13. A. flavus 656-2 was transformed with this construct. Aflatoxin production, GUS activity, and transcript accumulation were determined in transformants after shifting the cultures from a nonconducive medium to a medium conducive to aflatoxin biosynthesis. Transformants harboring GAP13 displayed GUS expression only when aflatoxin was detected in culture. Further, the transcription of the uidA gene driven by the ver1 promoter followed the same profile as for the ver1 genes. The results show that the GAP13 construct may be useful as a genetic tool to study the induction of aflatoxin in situ and to identify substances that affect the expression of genes involved in aflatoxin biosynthesis. The utility of this construct to detect inducers of aflatoxin biosynthesis in maize kernels was tested in a bioassay. A heat-stable inducer of aflatoxin with a molecular size of less than 10 kDa was detected in extracts from maize kernels colonized by A. flavus
ISSN:0099-2240
1098-5336
DOI:10.1128/aem.61.7.2482-2486.1995