Cargando…
Development of a transformation system for Aspergillus sojae based on the Agrobacterium tumefaciens-mediated approach
BACKGROUND: Aspergillus sojae has been an important filamentous fungus in Biotechnology due to its use in diverse fermentative processes for the production of various food products. Furthermore, this fungus is a common expression system for the production of enzymes and other metabolites. The availa...
Autores principales: | , , , |
---|---|
Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
BioMed Central
2014
|
Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4186950/ https://www.ncbi.nlm.nih.gov/pubmed/25253558 http://dx.doi.org/10.1186/s12866-014-0247-x |
_version_ | 1782338134928785408 |
---|---|
author | Mora-Lugo, Rodrigo Zimmermann, Judith Rizk, Amira M Fernandez-Lahore, Marcelo |
author_facet | Mora-Lugo, Rodrigo Zimmermann, Judith Rizk, Amira M Fernandez-Lahore, Marcelo |
author_sort | Mora-Lugo, Rodrigo |
collection | PubMed |
description | BACKGROUND: Aspergillus sojae has been an important filamentous fungus in Biotechnology due to its use in diverse fermentative processes for the production of various food products. Furthermore, this fungus is a common expression system for the production of enzymes and other metabolites. The availability of molecular genetic tools to explore its biology is thus of big interest. In this study, an Agrobacterium tumefaciens-mediated transformation (ATMT) system for A. sojae was developed and its applicability evaluated. RESULTS: The donor plasmid named pRM-eGFP was constructed for ATMT of A. sojae. This plasmid contains the ble and egfp genes in its transfer DNA element (T-DNA) to confer phleomycin resistance and express the enhanced green fluorescent protein (EGFP) in A. sojae, respectively. Agrobacterium tumefaciens (LBA4404) harboring the donor plasmid and A. sojae (ATCC 20235) were co-cultured under diverse conditions to achieve ATMT. The maximum number of transformed fungi was obtained after three days of co-culturing at 28°C, and selection with 50 μg/ml phleomycin. Polymerase chain reaction (PCR), fluorescence microscopy and Western Blot analysis for EGFP expression confirmed successful genomic integration of the T-DNA element in A. sojae. The T-DNA was mitotically stable in approximately 40% of the fungal transformants after four generations of sub-culturing under phleomycin pressure. CONCLUSION: We successfully established a new ATMT protocol for A. sojae. This transformation system should enable further protein expression studies on this filamentous fungus. |
format | Online Article Text |
id | pubmed-4186950 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2014 |
publisher | BioMed Central |
record_format | MEDLINE/PubMed |
spelling | pubmed-41869502014-10-08 Development of a transformation system for Aspergillus sojae based on the Agrobacterium tumefaciens-mediated approach Mora-Lugo, Rodrigo Zimmermann, Judith Rizk, Amira M Fernandez-Lahore, Marcelo BMC Microbiol Research Article BACKGROUND: Aspergillus sojae has been an important filamentous fungus in Biotechnology due to its use in diverse fermentative processes for the production of various food products. Furthermore, this fungus is a common expression system for the production of enzymes and other metabolites. The availability of molecular genetic tools to explore its biology is thus of big interest. In this study, an Agrobacterium tumefaciens-mediated transformation (ATMT) system for A. sojae was developed and its applicability evaluated. RESULTS: The donor plasmid named pRM-eGFP was constructed for ATMT of A. sojae. This plasmid contains the ble and egfp genes in its transfer DNA element (T-DNA) to confer phleomycin resistance and express the enhanced green fluorescent protein (EGFP) in A. sojae, respectively. Agrobacterium tumefaciens (LBA4404) harboring the donor plasmid and A. sojae (ATCC 20235) were co-cultured under diverse conditions to achieve ATMT. The maximum number of transformed fungi was obtained after three days of co-culturing at 28°C, and selection with 50 μg/ml phleomycin. Polymerase chain reaction (PCR), fluorescence microscopy and Western Blot analysis for EGFP expression confirmed successful genomic integration of the T-DNA element in A. sojae. The T-DNA was mitotically stable in approximately 40% of the fungal transformants after four generations of sub-culturing under phleomycin pressure. CONCLUSION: We successfully established a new ATMT protocol for A. sojae. This transformation system should enable further protein expression studies on this filamentous fungus. BioMed Central 2014-09-25 /pmc/articles/PMC4186950/ /pubmed/25253558 http://dx.doi.org/10.1186/s12866-014-0247-x Text en © Mora-Lugo et al.; licensee BioMed Central Ltd. 2014 This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/4.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly credited. The Creative Commons Public Domain Dedication waiver (http://creativecommons.org/publicdomain/zero/1.0/) applies to the data made available in this article, unless otherwise stated. |
spellingShingle | Research Article Mora-Lugo, Rodrigo Zimmermann, Judith Rizk, Amira M Fernandez-Lahore, Marcelo Development of a transformation system for Aspergillus sojae based on the Agrobacterium tumefaciens-mediated approach |
title | Development of a transformation system for Aspergillus sojae based on the Agrobacterium tumefaciens-mediated approach |
title_full | Development of a transformation system for Aspergillus sojae based on the Agrobacterium tumefaciens-mediated approach |
title_fullStr | Development of a transformation system for Aspergillus sojae based on the Agrobacterium tumefaciens-mediated approach |
title_full_unstemmed | Development of a transformation system for Aspergillus sojae based on the Agrobacterium tumefaciens-mediated approach |
title_short | Development of a transformation system for Aspergillus sojae based on the Agrobacterium tumefaciens-mediated approach |
title_sort | development of a transformation system for aspergillus sojae based on the agrobacterium tumefaciens-mediated approach |
topic | Research Article |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4186950/ https://www.ncbi.nlm.nih.gov/pubmed/25253558 http://dx.doi.org/10.1186/s12866-014-0247-x |
work_keys_str_mv | AT moralugorodrigo developmentofatransformationsystemforaspergillussojaebasedontheagrobacteriumtumefaciensmediatedapproach AT zimmermannjudith developmentofatransformationsystemforaspergillussojaebasedontheagrobacteriumtumefaciensmediatedapproach AT rizkamiram developmentofatransformationsystemforaspergillussojaebasedontheagrobacteriumtumefaciensmediatedapproach AT fernandezlahoremarcelo developmentofatransformationsystemforaspergillussojaebasedontheagrobacteriumtumefaciensmediatedapproach |