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Gateway-compatible vectors for functional analysis of proteins in cell type specific manner

BACKGROUND: Genetically encoded fluorescent proteins are often used to label proteins and study protein function and localization in vivo. Traditional cloning methods mediated by restriction digestion and ligation are time-consuming and sometimes difficult due to the lack of suitable restriction sit...

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Autores principales: Zhang, Liu, Zhao, Yang, Liang, Haiyan, Li, Xugang, Gallagher, Kimberly L., Wu, Shuang
Formato: Online Artículo Texto
Lenguaje:English
Publicado: BioMed Central 2020
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7339564/
https://www.ncbi.nlm.nih.gov/pubmed/32655679
http://dx.doi.org/10.1186/s13007-020-00635-z
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author Zhang, Liu
Zhao, Yang
Liang, Haiyan
Li, Xugang
Gallagher, Kimberly L.
Wu, Shuang
author_facet Zhang, Liu
Zhao, Yang
Liang, Haiyan
Li, Xugang
Gallagher, Kimberly L.
Wu, Shuang
author_sort Zhang, Liu
collection PubMed
description BACKGROUND: Genetically encoded fluorescent proteins are often used to label proteins and study protein function and localization in vivo. Traditional cloning methods mediated by restriction digestion and ligation are time-consuming and sometimes difficult due to the lack of suitable restriction sites. Invitrogen developed the Gateway cloning system based on the site-specific DNA recombination, which allows for digestion-free cloning. Most gateway destination vectors available for use in plants employ either the 35S or ubiquitin promoters, which confer high-level, ubiquitous expression. There are far fewer options for moderate, cell-type specific expression. RESULTS: Here we report on the construction of a Gateway-compatible cloning system (SWU vectors) to rapidly tag various proteins and express them in a cell-type specific manner in plants. We tested the SWU vectors using the HISTONE (H2B) coding sequence in stable transgenic plants. CONCLUSIONS: The SWU vectors are a valuable tool for low cost, high efficiency functional analysis of proteins of interest in specific cell types in the Arabidopsis root.
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spelling pubmed-73395642020-07-09 Gateway-compatible vectors for functional analysis of proteins in cell type specific manner Zhang, Liu Zhao, Yang Liang, Haiyan Li, Xugang Gallagher, Kimberly L. Wu, Shuang Plant Methods Methodology BACKGROUND: Genetically encoded fluorescent proteins are often used to label proteins and study protein function and localization in vivo. Traditional cloning methods mediated by restriction digestion and ligation are time-consuming and sometimes difficult due to the lack of suitable restriction sites. Invitrogen developed the Gateway cloning system based on the site-specific DNA recombination, which allows for digestion-free cloning. Most gateway destination vectors available for use in plants employ either the 35S or ubiquitin promoters, which confer high-level, ubiquitous expression. There are far fewer options for moderate, cell-type specific expression. RESULTS: Here we report on the construction of a Gateway-compatible cloning system (SWU vectors) to rapidly tag various proteins and express them in a cell-type specific manner in plants. We tested the SWU vectors using the HISTONE (H2B) coding sequence in stable transgenic plants. CONCLUSIONS: The SWU vectors are a valuable tool for low cost, high efficiency functional analysis of proteins of interest in specific cell types in the Arabidopsis root. BioMed Central 2020-07-06 /pmc/articles/PMC7339564/ /pubmed/32655679 http://dx.doi.org/10.1186/s13007-020-00635-z Text en © The Author(s) 2020 Open AccessThis article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons licence, and indicate if changes were made. The images or other third party material in this article are included in the article's Creative Commons licence, unless indicated otherwise in a credit line to the material. If material is not included in the article's Creative Commons licence and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this licence, visit http://creativecommons.org/licenses/by/4.0/. 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 in a credit line to the data.
spellingShingle Methodology
Zhang, Liu
Zhao, Yang
Liang, Haiyan
Li, Xugang
Gallagher, Kimberly L.
Wu, Shuang
Gateway-compatible vectors for functional analysis of proteins in cell type specific manner
title Gateway-compatible vectors for functional analysis of proteins in cell type specific manner
title_full Gateway-compatible vectors for functional analysis of proteins in cell type specific manner
title_fullStr Gateway-compatible vectors for functional analysis of proteins in cell type specific manner
title_full_unstemmed Gateway-compatible vectors for functional analysis of proteins in cell type specific manner
title_short Gateway-compatible vectors for functional analysis of proteins in cell type specific manner
title_sort gateway-compatible vectors for functional analysis of proteins in cell type specific manner
topic Methodology
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC7339564/
https://www.ncbi.nlm.nih.gov/pubmed/32655679
http://dx.doi.org/10.1186/s13007-020-00635-z
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