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Particle bombardment and subcellular protein localization analysis in the aquatic plant Egeria densa
Particle bombardment is a powerful and relatively easy method for transient expression of genes of interest in plant cells, especially those that are recalcitrant to other transformation methods. This method has facilitated numerous analyses of subcellular localization of fluorescent fusion protein...
Autores principales: | , |
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Formato: | Online Artículo Texto |
Lenguaje: | English |
Publicado: |
PeerJ Inc.
2017
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Materias: | |
Acceso en línea: | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5592081/ https://www.ncbi.nlm.nih.gov/pubmed/28894649 http://dx.doi.org/10.7717/peerj.3779 |
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author | Osaki, Yasuhide Kodama, Yutaka |
author_facet | Osaki, Yasuhide Kodama, Yutaka |
author_sort | Osaki, Yasuhide |
collection | PubMed |
description | Particle bombardment is a powerful and relatively easy method for transient expression of genes of interest in plant cells, especially those that are recalcitrant to other transformation methods. This method has facilitated numerous analyses of subcellular localization of fluorescent fusion protein constructs. Particle bombardment delivers genes to the first layer of plant tissue. In leaves of higher plants, epidermal cells are the first cell layer. Many studies have used the epidermal cell layer of onion bulb (Allium cepa) as the experimental tissue, because these cells are relatively large. However, onion epidermal cells lack developed plastids (i.e., chloroplasts), thereby precluding subcellular localization analysis of chloroplastic proteins. In this study, we developed a protocol for particle bombardment of the aquatic plant Egeria densa, and showed that it is a useful system for subcellular localization analysis of higher plant proteins. E. densa leaflets contain only two cell layers, and cells in the adaxial layer are sufficiently large for observation. The cells in both layers contain well-developed chloroplasts. We fused fluorescent proteins to conventional plant localization signals for the nucleus, cytosol, mitochondria, peroxisome, and chloroplast, and used particle bombardment to transiently express these fusion constructs in E. densa leaves. The plant subcellular localization signals functioned normally and displayed the expected distributions in transiently transformed E. densa cells, and even chloroplastic structures could be clearly visualized. |
format | Online Article Text |
id | pubmed-5592081 |
institution | National Center for Biotechnology Information |
language | English |
publishDate | 2017 |
publisher | PeerJ Inc. |
record_format | MEDLINE/PubMed |
spelling | pubmed-55920812017-09-11 Particle bombardment and subcellular protein localization analysis in the aquatic plant Egeria densa Osaki, Yasuhide Kodama, Yutaka PeerJ Cell Biology Particle bombardment is a powerful and relatively easy method for transient expression of genes of interest in plant cells, especially those that are recalcitrant to other transformation methods. This method has facilitated numerous analyses of subcellular localization of fluorescent fusion protein constructs. Particle bombardment delivers genes to the first layer of plant tissue. In leaves of higher plants, epidermal cells are the first cell layer. Many studies have used the epidermal cell layer of onion bulb (Allium cepa) as the experimental tissue, because these cells are relatively large. However, onion epidermal cells lack developed plastids (i.e., chloroplasts), thereby precluding subcellular localization analysis of chloroplastic proteins. In this study, we developed a protocol for particle bombardment of the aquatic plant Egeria densa, and showed that it is a useful system for subcellular localization analysis of higher plant proteins. E. densa leaflets contain only two cell layers, and cells in the adaxial layer are sufficiently large for observation. The cells in both layers contain well-developed chloroplasts. We fused fluorescent proteins to conventional plant localization signals for the nucleus, cytosol, mitochondria, peroxisome, and chloroplast, and used particle bombardment to transiently express these fusion constructs in E. densa leaves. The plant subcellular localization signals functioned normally and displayed the expected distributions in transiently transformed E. densa cells, and even chloroplastic structures could be clearly visualized. PeerJ Inc. 2017-09-07 /pmc/articles/PMC5592081/ /pubmed/28894649 http://dx.doi.org/10.7717/peerj.3779 Text en ©2017 Osaki and Kodama http://creativecommons.org/licenses/by/4.0/ 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, reproduction and adaptation in any medium and for any purpose provided that it is properly attributed. For attribution, the original author(s), title, publication source (PeerJ) and either DOI or URL of the article must be cited. |
spellingShingle | Cell Biology Osaki, Yasuhide Kodama, Yutaka Particle bombardment and subcellular protein localization analysis in the aquatic plant Egeria densa |
title | Particle bombardment and subcellular protein localization analysis in the aquatic plant Egeria densa |
title_full | Particle bombardment and subcellular protein localization analysis in the aquatic plant Egeria densa |
title_fullStr | Particle bombardment and subcellular protein localization analysis in the aquatic plant Egeria densa |
title_full_unstemmed | Particle bombardment and subcellular protein localization analysis in the aquatic plant Egeria densa |
title_short | Particle bombardment and subcellular protein localization analysis in the aquatic plant Egeria densa |
title_sort | particle bombardment and subcellular protein localization analysis in the aquatic plant egeria densa |
topic | Cell Biology |
url | https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5592081/ https://www.ncbi.nlm.nih.gov/pubmed/28894649 http://dx.doi.org/10.7717/peerj.3779 |
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