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Cloning, Overexpression and in vitro Antifungal Activity of Zea Mays PR10 Protein
BACKGROUND: Plants have various defense mechanisms such as production of antimicrobial peptides, particularly pathogenesis related proteins (PR proteins). PR10 family is an essential member of this group, with antifungal, antibacterial and antiviral activities. OBJECTIVE: The goal of this study is t...
Autores principales: | , , , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
National Institute of Genetic Engineering and Biotechnology
2017
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Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5582252/ https://www.ncbi.nlm.nih.gov/pubmed/28959351 http://dx.doi.org/10.15171/ijb.1357 |
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author | Zandvakili, Niloofar Zamani, Mohammadreza Motallebi, Mostafa Moghaddassi Jahromi, Zahra |
author_facet | Zandvakili, Niloofar Zamani, Mohammadreza Motallebi, Mostafa Moghaddassi Jahromi, Zahra |
author_sort | Zandvakili, Niloofar |
collection | PubMed |
description | BACKGROUND: Plants have various defense mechanisms such as production of antimicrobial peptides, particularly pathogenesis related proteins (PR proteins). PR10 family is an essential member of this group, with antifungal, antibacterial and antiviral activities. OBJECTIVE: The goal of this study is to assess the antifungal activity of maize PR10 against some of fungal phytopathogens. MATERIALS AND METHODS: Zea mays PR10 gene (TN-05-147) was cloned from genomic DNA and cDNA and overexpressed in Escherichia coli. The existence of a 77- bp intron and two exons in PR10 was confi rmed by comparing the genomic and cDNA sequences. The PR10 cDNA was cloned in pET26b (+) expression vector and transformed into E. coli strain Rosetta DE3 in order to express PR10 recombinant protein. Expression of the recombinant protein was checked by western analysis. Recombinant PR10 appeared as insoluble inclusion bodies and thus solubilized and refolded. PR10 was isolated using Ni- NTA column. The activity of the refolded protein was confi rmed by DNA degradation test. The antifungal activity of PR10 was assessed using radial diff usion, disc diff usion and spore germination. The hemolytic assay was performed to investigate the biosafety of recombinant PR10. RESULTS: Recombinant maize PR10 exerted broad spectrum antifungal activity against Botrytis cinerea, Sclerotinia sclerotiorum, Fusarium oxysporum, Verticillium dahlia and Alternaria solani. Hemolysis biosafety test indicated that the protein is not poisonous to mammalian cells. CONCLUSIONS: Maize PR10 has the potential to be used as the antifungal agent against diff erent fungal phytopathogens. Therefore, this protein can be used in order to produce antifungal agents and fungi resistance transgenic plants. |
format | Online Article Text |
id | pubmed-5582252 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2017 |
publisher | National Institute of Genetic Engineering and Biotechnology |
record_format | MEDLINE/PubMed |
spelling | pubmed-55822522017-09-28 Cloning, Overexpression and in vitro Antifungal Activity of Zea Mays PR10 Protein Zandvakili, Niloofar Zamani, Mohammadreza Motallebi, Mostafa Moghaddassi Jahromi, Zahra Iran J Biotechnol Research Article BACKGROUND: Plants have various defense mechanisms such as production of antimicrobial peptides, particularly pathogenesis related proteins (PR proteins). PR10 family is an essential member of this group, with antifungal, antibacterial and antiviral activities. OBJECTIVE: The goal of this study is to assess the antifungal activity of maize PR10 against some of fungal phytopathogens. MATERIALS AND METHODS: Zea mays PR10 gene (TN-05-147) was cloned from genomic DNA and cDNA and overexpressed in Escherichia coli. The existence of a 77- bp intron and two exons in PR10 was confi rmed by comparing the genomic and cDNA sequences. The PR10 cDNA was cloned in pET26b (+) expression vector and transformed into E. coli strain Rosetta DE3 in order to express PR10 recombinant protein. Expression of the recombinant protein was checked by western analysis. Recombinant PR10 appeared as insoluble inclusion bodies and thus solubilized and refolded. PR10 was isolated using Ni- NTA column. The activity of the refolded protein was confi rmed by DNA degradation test. The antifungal activity of PR10 was assessed using radial diff usion, disc diff usion and spore germination. The hemolytic assay was performed to investigate the biosafety of recombinant PR10. RESULTS: Recombinant maize PR10 exerted broad spectrum antifungal activity against Botrytis cinerea, Sclerotinia sclerotiorum, Fusarium oxysporum, Verticillium dahlia and Alternaria solani. Hemolysis biosafety test indicated that the protein is not poisonous to mammalian cells. CONCLUSIONS: Maize PR10 has the potential to be used as the antifungal agent against diff erent fungal phytopathogens. Therefore, this protein can be used in order to produce antifungal agents and fungi resistance transgenic plants. National Institute of Genetic Engineering and Biotechnology 2017-03 /pmc/articles/PMC5582252/ /pubmed/28959351 http://dx.doi.org/10.15171/ijb.1357 Text en © 2017 by National Institute of Genetic Engineering and Biotechnology https://creativecommons.org/licenses/by/4.0/ This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. |
spellingShingle | Research Article Zandvakili, Niloofar Zamani, Mohammadreza Motallebi, Mostafa Moghaddassi Jahromi, Zahra Cloning, Overexpression and in vitro Antifungal Activity of Zea Mays PR10 Protein |
title |
Cloning, Overexpression and in vitro Antifungal Activity of Zea Mays PR10 Protein
|
title_full |
Cloning, Overexpression and in vitro Antifungal Activity of Zea Mays PR10 Protein
|
title_fullStr |
Cloning, Overexpression and in vitro Antifungal Activity of Zea Mays PR10 Protein
|
title_full_unstemmed |
Cloning, Overexpression and in vitro Antifungal Activity of Zea Mays PR10 Protein
|
title_short |
Cloning, Overexpression and in vitro Antifungal Activity of Zea Mays PR10 Protein
|
title_sort | cloning, overexpression and in vitro antifungal activity of zea mays pr10 protein |
topic | Research Article |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5582252/ https://www.ncbi.nlm.nih.gov/pubmed/28959351 http://dx.doi.org/10.15171/ijb.1357 |
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