Please use this identifier to cite or link to this item: http://dx.doi.org/10.14279/depositonce-7051
Main Title: HisB as novel selection marker for gene targeting approaches in Aspergillus niger
Author(s): Fiedler, Markus R. M.
Gensheimer, Tarek
Kubisch, Christin
Meyer, Vera
Type: Article
Language Code: en
Abstract: Background For Aspergillus niger, a broad set of auxotrophic and dominant resistance markers is available. However, only few offer targeted modification of a gene of interest into or at a genomic locus of choice, which hampers functional genomics studies. We thus aimed to extend the available set by generating a histidine auxotrophic strain with a characterized hisB locus for targeted gene integration and deletion in A. niger. Results A histidine-auxotrophic strain was established via disruption of the A. niger hisB gene by using the counterselectable pyrG marker. After curing, a hisB - , pyrG - strain was obtained, which served as recipient strain for further studies. We show here that both hisB orthologs from A. nidulans and A. niger can be used to reestablish histidine prototrophy in this recipient strain. Whereas the hisB gene from A. nidulans was suitable for efficient gene targeting at different loci in A. niger, the hisB gene from A. niger allowed efficient integration of a Tet-on driven luciferase reporter construct at the endogenous non-functional hisB locus. Subsequent analysis of the luciferase activity revealed that the hisB locus is tight under non-inducing conditions and allows even higher luciferase expression levels compared to the pyrG integration locus. Conclusion Taken together, we provide here an alternative selection marker for A. niger, hisB, which allows efficient homologous integration rates as well as high expression levels which compare favorably to the well-established pyrG selection marker.
URI: https://depositonce.tu-berlin.de//handle/11303/7890
http://dx.doi.org/10.14279/depositonce-7051
Issue Date: 8-Mar-2017
Date Available: 31-May-2018
DDC Class: 570 Biowissenschaften; Biologie
Subject(s): Aspergillus niger
gene expression
selection marker
HisB
Tet-On
Sponsor/Funder: EC/FP7/303864/EU/Bridging the world of fungi and dementia/
License: https://creativecommons.org/licenses/by/4.0/
Journal Title: BMC Microbiology
Publisher: BioMed Central
Publisher Place: London
Volume: 17
Article Number: 57
Publisher DOI: 10.1186/s12866-017-0960-3
ISSN: 1471-2180
Appears in Collections:FG Angewandte und Molekulare Mikrobiologie » Publications

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