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The Aspergillus nidulans amdS gene as a marker for the identification of multicopy T-DNA integration events in Agrobacterium-mediated transformation of Aspergillus awamori

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Author: Michielse, C.B. · Ram, A.F.J. · Hondel, C.A.M.J.J. van den
Type:article
Date:2004
Institution: TNO Voeding
Source:Current Genetics, 6, 45, 399-403
Identifier: 237834
doi: doi:10.1007/s00294-004-0500-1
Keywords: Biology · Biotechnology · Agrobacterium tumefaciens · amdS selection marker · Aspergillus awamori · Filamentous fungi · Multicopy T-DNA integration · Transformation · acetamidase · acetamide · agar · DNA · fungal enzyme · hygromycin · nitrogen · unclassified drug · agar medium · analytic method · antibiotic resistance · article · Aspergillus awamori · Aspergillus nidulans · controlled study · culture medium · DNA integration · frequency analysis · fungus identification · gene transfer · genetic selection · genetic transformation · growth rate · intermethod comparison · marker gene · nonhuman · priority journal · Rhizobium · Amidohydrolases · Aspergillus · Cinnamates · Gene Expression Regulation, Fungal · Genes, Fungal · Genetic Markers · Hygromycin B · Plasmids · Rhizobium · Transformation, Genetic · Agrobacterium · Agrobacterium tumefaciens · Aspergillus · Aspergillus awamori · Emericella nidulans · Fungi

Abstract

The Aspergillus nidulans amdS selection marker was used for the identification of multicopy T-DNA insertions in Agrobacterium-mediated transformation of Asp. awamori. The selection of transformants on agar plates containing acetamide as sole nitrogen source and hygromycin resulted in a six-fold decrease in the transformation frequency, compared with the transformation frequency obtained after hygromycin selection alone. However, it was found that 47% of the transformants obtained after hygromycin and acetamide double selection contained multiple T-DNA integrations. Furthermore, it was found that the multicopy transformants could easily be identified based on their growth rate on agar plates containing acetamide medium. Based on these data, it can be concluded that the amdS marker can also be used as a selection marker in Agrobacterium-mediated transformation of Asp. awamori and that it is a very useful marker to identify those transformants containing multiple T-DNA integrations. © Springer-Verlag 2004.